OBJECTIVE: Here we describe a novel IncFIA plasmid harbouring mcr-10 gene in a clinical Enterobacter ludwigii strain isolated at the University Hospital in Pilsen in the Czech Republic. METHODS: The strain was subjected to antibiotic susceptibility testing. Whole genome sequencing was performed using Illumina for short-read sequencing and Oxford Nanopore Technologies for long-read sequencing followed by hybrid assembly. The resulting genome was used to detect species using average nucleotide identity, resistance genes, plasmid replicon and MLST (using centre for genomic epidemiology databases; ResFinder, PlasmidFinder and MLST, respectively) and virulence genes using VFDB. RESULTS: Τhe strain showed susceptibility against tetracycline, cefuroxime and chloramphenicol, and it was susceptible to the second and third generation of cephalosporins, carbapenems and colistin. Genome analysis identified the strain as E. ludwigii sequence type ST20 and located the mcr-10 gene on an IncFIA (HI1)/IncFII (Yp) plasmid (pI9455333_MCR10; 129 863 bp). Upon blasting the nucleotide sequence of pI9455333_MCR10 against the NCBI database, no similar plasmid sequence was detected, implying a novel plasmid structure. Nevertheless, it showed a partial similarity with pRHBSTW-00123_3 and FDAARGOS 1432, which were detected in Enterobacter cloacae complex (ECC) strains in wastewater samples in 2017 in UK and in 2021 in the United States, respectively, and pEC81-mcr, which was detected in a clinical Escherichia coli strain in 2020 in China. Moreover, I9455333cz genome carried virulence genes coding for curli fibers, fimbrial adherence determinants, siderophore aerobactin, iron uptake proteins and regulators of sigma factor. CONCLUSION: In conclusion, we identified a novel IncF plasmid harbouring mcr-10 gene in a clinical Enterobacter ludwigii strain. To our knowledge, this is the first clinical report of mcr-10 in the Czech Republic.
- MeSH
- antibakteriální látky * farmakologie MeSH
- bakteriální proteiny genetika MeSH
- centra terciární péče * MeSH
- Enterobacter * genetika účinky léků izolace a purifikace MeSH
- enterobakteriální infekce * mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti * MeSH
- mnohočetná bakteriální léková rezistence genetika MeSH
- multilokusová sekvenční typizace MeSH
- plazmidy * genetika MeSH
- sekvenování celého genomu MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
Alginate lyases have countless potential for application in industries and medicine particularly as an appealing biocatalyst for the production of biofuels and bioactive oligosaccharides. Solid-state fermentation (SSF) allows improved production of enzymes and consumes less energy compared to submerged fermentation. Seaweeds can serve as the most promising biomass for the production of biochemicals. Alginate present in the seaweed can be used by alginate lyase-producing bacteria to support growth and can secrete alginate lyase. In this perspective, the current study was directed on the bioprocessing of brown seaweeds for the production of alginate lyase using marine bacterial isolate. A novel alginate-degrading marine bacterium Enterobacter tabaci RAU2C which was previously isolated in the laboratory was used for the production of alginate lyase using Sargassum swartzii as a low-cost solid substrate. Process parameters such as inoculum incubation period and moisture content were optimized for alginate lyase production. SSF resulted in 33.56 U/mL of alginate lyase under the static condition maintained with 75% moisture after 4 days. Further, the effect of different buffers, pH, and temperature on alginate lyase activity was also analyzed. An increase in alginate lyase activity was observed with an increase in moisture content from 60 to 75%. Maximum enzyme activity was perceived with phosphate buffer at pH 7 and 37 °C. Further, the residual biomass after SSF could be employed as biofertilizer for plant growth promotion based on the preliminary analysis. To our knowledge, this is the first report stating the usage of seaweed biomass as a substrate for the production of alginate lyase using solid-state fermentation.
- MeSH
- algináty * metabolismus MeSH
- biomasa MeSH
- Enterobacter * metabolismus enzymologie izolace a purifikace růst a vývoj MeSH
- fermentace * MeSH
- koncentrace vodíkových iontů MeSH
- kyselina glukuronová metabolismus MeSH
- mořské řasy * mikrobiologie MeSH
- Phaeophyceae mikrobiologie MeSH
- polysacharid-lyasy * metabolismus MeSH
- Sargassum * mikrobiologie metabolismus MeSH
- teplota MeSH
- Publikační typ
- časopisecké články MeSH
Reducing bacterial pathogen contamination not only improves overall global public health but also diminishes food waste and loss. The use of lytic bacteriophages (phages) that infect and kill bacteria could be a beneficial tool for suppressing bacterial growth during dairy products storage time. Four Enterobacter cloacae (E. cloacae) complex isolates which were previously isolated from contaminated dairy products were used to identify lytic phages in wastewater. Phages specific to multi-drug resistant (MDR) E. cloacae complex 6AS1 were isolated from local sewage. Two novel phages vB_EclM-EP1 and vB_EclM-EP2 were identified as myoviral particles and have double-stranded DNA genome. Their host range and lytic capabilities were detected using spot test and efficiency of plating (EOP) against several bacterial isolates. The phages had a latent period of 30 min, and a large burst size of about 100 and 142 PFU/cell for vB_EclM-EP1 and vB_EclM-EP2, respectively. Both phages were viable at pH ranging 5-9 and stable at 70 °C for 60 min. The individual phages and their cocktail preparations (vB_EclM-EP1 and vB_EclM-EP2) reduced and inhibited the growth of E. cloacae complex 6AS1 during challenge test in milk and yogurt samples. These results indicate that the E. cloacae complex-specific phages (vB_EclM-EP1 and vB_EclM-EP2) have a potential application as microbicidal agents in packaged milk and milk derivatives during storage time. In addition, our environment is a rich sources of lytic phages which have potential use in eliminating multidrug-resistant isolates in food industry as well as in biocontrol.
- MeSH
- bakteriofágy * genetika MeSH
- Enterobacter cloacae MeSH
- jogurt MeSH
- mléko mikrobiologie MeSH
- odpadky - odstraňování * MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Enterobacter spp. and Klebsiella aerogenes are rod-shaped Gram-negative opportunistic pathogens. This study aimed at the molecular and genomic characterization of multidrug resistant Enterobacter spp. and K. aerogenes isolates recovered from hospitalized patients in a tertiary care hospital in Lebanon. A total of 59 Enterobacter spp. clinical isolates consisting of 41 carbapenem-resistant and 18 susceptible by Etest were included in this study. Genotypic identification through whole-genome sequencing (WGS) was performed and confirmed in silico. Resistance and plasmid profiles were studied using ResFinder4.0 and Plasmid-Finder2.1. Multilocus sequence typing (MLST) was used to determine the isolates' clonality. Using the average nucleotide identity (ANI) we identified and confirmed that 47 (80%) isolates were E. hormaechei, 11 (18%) were Klebsiella aerogenes and 1 (2%) was an E. cloacae. Carbapenem-resistance was detected among 41 isolates all showing an MIC90 of ≥ 32 μg/mL for ertapenem, imipenem, and meropenem. blaNDM-1 (58.5%), blaACT-16 (54%), and blaOXA-1 (54%) were the most common detected β-lactamases, while blaCTX-M-15 (68%) was the main detected extended-spectrum β-lactamase (ESBL) encoding gene. Chromosomal ampC, carbapenemase encoding genes, and porin modifications were among the detected carbapenem resistance determinants. The carbapenemase encoding genes were linked to three well-defined plasmid Inc groups, IncFII/IncFIB, IncX3, and IncL. MLST typing revealed the diversity within the studied isolates, with ST114 being the most common among the studied E. hormaechei.: The spread of carbapenem-resistant isolates in clinical settings in Lebanon is a serious challenge. Screening and continuous monitoring through WGS analysis could effectively limit the dissemination of drug-resistant isolates in hospitalized patients. IMPORTANCE Drug resistance is an increasing global public health threat that involves most disease-causing organisms and antimicrobial drugs. Drug-resistant organisms spread in health care settings, and resistance to multiple drugs is common. Our study demonstrated the mechanisms leading to resistance against the last resort antimicrobial agents among members of the Enterobacteriaceae family. The spread of carbapenem-resistant bacteria in clinical settings is a serious challenge. Screening and continuous monitoring could effectively limit the dissemination of drug-resistant isolates in hospitalized patients.
- MeSH
- antibakteriální látky farmakologie terapeutické užití MeSH
- bakteriální proteiny genetika MeSH
- beta-laktamasy genetika MeSH
- Enterobacter aerogenes * genetika MeSH
- Enterobacter genetika MeSH
- karbapenemy farmakologie MeSH
- Klebsiella pneumoniae genetika MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- multilokusová sekvenční typizace MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Libanon MeSH
In the present study, bacterial isolates were screened for arsenic resistance efficiency. Environmental isolates were isolated from arsenic-rich soil samples (i.e., from Rajnandgaon district of Chhattisgarh state, India). Amplification and sequencing of 16S rRNA gene revealed that the isolates were of Bacillus firmus RSN1, Brevibacterium senegalense RSN2, Enterobacter cloacae RSN3, Stenotrophomonas pavanii RSN6, Achromobacter mucicolens RSN7, and Ochrobactrum intermedium RSN10. Arsenite efflux gene (arsB) was successfully amplified in E. cloacae RSN3. Atomic absorption spectroscopy (AAS) analysis showed an absorption of 32.22% arsenic by the RSN3 strain. Furthermore, results of scanning electron microscopy (SEM) for morphological variations revealed an initial increase in the cell size at 1 mM sodium arsenate; however, it was decreased at 10 mM concentration in comparison to control. This change of the cell size in different metal concentrations was due to the uptake and expulsion of the metal from the cell, which also confirmed the arsenite efflux system.
Infections caused by Methicillin-Resistant Staphylococcus aureus (MRSA) and Extended-Spectrum Beta-Lactamase (ESBL) producing Enterobacter cloacae are considered as major therapeutic challenge due to their multidrug-resistant (MDR) phenotype against conventional antibiotics. WLBU2 is an engineered cationic peptide with potent antimicrobial activity. This in-vitro study aimed to evaluate the effects of WLBU2 against clinical isolates of the aforementioned bacteria and assess whether synergistic effects can be achieved upon combination with conventional antibiotics. The minimum inhibitory concentrations (MICs) of antimicrobial agents against bacterial clinical isolates (n = 30/strain) were determined using the microbroth dilution assay. The minimum bactericidal concentrations (MBCs) of WLBU2 were determined from microbroth dilution (MICs) tests by subculturing to agar plates. MICs of WLBU2 were evaluated in the presence of physiological concentrations of salts including NaCl, CaCl2 and MgCl2. To identify bacterial resistance profile, MRSA were treated with Oxacillin, Erythromycin and Vancomycin, while Ceftazidime, Ceftriaxone, Ciprofloxacin and Imipenem were used against Enterobacter cloacae. Combination treatments of antibiotics and sub-inhibitory concentrations of WLBU2 were conducted when MICs indicated intermediate/resistant susceptibility. The MICs/MBCs of WLBU2 were identical for each respective bacteria with values of 0.78-6.25 μM and 1.5-12.5 μM against MRSA and Enterobacter cloacae, respectively. WLBU2 was found as salt resistant. Combination treatment showed that synergistic and additive effects were achieved in many isolates of MRSA and Enterobacter cloacae. Our data revealed that WLBU2 is a potent peptide with bactericidal activity. In addition, it demonstrated the selective advantage of WLBU2 as a potential therapeutic agent under physiological solutions. Our findings also support the combination of WLBU2 and conventional antibiotics with potential application for treatment of resistant bacteria.
- MeSH
- bakteriální léková rezistence MeSH
- Enterobacter cloacae * účinky léků MeSH
- kationické antimikrobiální peptidy farmakologie terapeutické užití MeSH
- kombinovaná farmakoterapie MeSH
- lidé MeSH
- Staphylococcus aureus * účinky léků MeSH
- synergismus léků * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Wild corvids were examined for the presence of carbapenemase-producing Gram-negative bacteria in the United States. A total of 13 isolates were detected among 590 fecal samples of American crow; 11 Providencia rettgeri isolates harboring blaIMP-27 on the chromosome as a class 2 integron gene cassette within the Tn7 transposon, 1 Klebsiella pneumoniae ST258 isolate carrying blaKPC-2 on a pKpQIL-like plasmid as a part of Tn4401a, and 1 Enterobacter bugandensis isolate with blaIMI-1 located within EcloIMEX-2.
- MeSH
- antibakteriální látky farmakologie MeSH
- bakteriální proteiny genetika MeSH
- beta-laktamasy genetika MeSH
- Enterobacter MeSH
- infekce bakteriemi rodu Klebsiella * MeSH
- Klebsiella pneumoniae genetika MeSH
- mikrobiální testy citlivosti MeSH
- plazmidy genetika MeSH
- Providencia MeSH
- vrány * MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Spojené státy americké MeSH
The aim of this study was to report the characterization of the first mcr-positive Enterobacterales isolated from Czech hospitals. In 2019, one Citrobacter freundii and four Enterobacter isolates were recovered from Czech hospitals. The production of carbapenemases was examined by a matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) imipenem hydrolysis assay. Additionally, bacteria were screened for the presence of carbapenemase-encoding genes and plasmid-mediated colistin resistance genes by PCR. To define the genetic units carrying mcr genes, the genomic DNAs of mcr-carrying clinical isolates were sequenced on the PacBio Sequel I platform. Results showed that all isolates carried blaVIM- and mcr-like genes. Analysis of whole-genome sequencing (WGS) data revealed that all isolates carried mcr-9-like alleles. Furthermore, the three sequence type 106 (ST106) Enterobacter hormaechei isolates harbored the blaVIM-1 gene, while the ST764 E. hormaechei and ST95 C. freundii included blaVIM-4 Analysis of plasmid sequences showed that, in all isolates, mcr-9 was carried on IncHI2 plasmids. Additionally, at least one multidrug resistance (MDR) region was identified in each mcr-9-carrying IncHI2 plasmid. The blaVIM-4 gene was found in the MDR regions of p48880_MCR_VIM and p51929_MCR_VIM. In the three remaining isolates, blaVIM-1 was localized on plasmids (∼55 kb) exhibiting repA-like sequences 99% identical to the respective gene of pKPC-CAV1193. In conclusion, to the best of our knowledge, these 5 isolates were the first mcr-9-positive bacteria of clinical origin identified in the Czech Republic. Additionally, the carriage of the blaVIM-1 on pKPC-CAV1193-like plasmids is described for the first time. Thus, our findings underline the ongoing evolution of mobile elements implicated in the dissemination of clinically important resistance determinants.IMPORTANCE Infections caused by carbapenemase-producing bacteria have led to the revival of polymyxins as the "last-resort" antibiotic. Since 2016, several reports describing the presence of plasmid-mediated colistin resistance genes, mcr, in different host species and geographic areas were published. Here, we report the first detection of Enterobacterales carrying mcr-9-like alleles isolated from Czech hospitals in 2019. Furthermore, the three ST106 Enterobacter hormaechei isolates harbored blaVIM-1, while the ST764 E. hormaechei and ST95 Citrobacter freundii isolates included blaVIM-4 Analysis of WGS data showed that, in all isolates, mcr-9 was carried on IncHI2 plasmids. blaVIM-4 was found in the MDR regions of IncHI2 plasmids, while blaVIM-1 was localized on pKPC-CAV1193-like plasmids, described here for the first time. These findings underline the ongoing evolution of mobile elements implicated in dissemination of clinically important resistance determinants. Thus, WGS characterization of MDR bacteria is crucial to unravel the mechanisms involved in dissemination of resistance mechanisms.
- MeSH
- antibakteriální látky farmakologie terapeutické užití MeSH
- bakteriální proteiny genetika MeSH
- beta-laktamasy genetika MeSH
- Enterobacter genetika izolace a purifikace MeSH
- lidé MeSH
- nemocnice MeSH
- plazmidy genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- multicentrická studie MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
Out of six samples of wastewater produced in the dairy industry, taken in 2017 at various places of dairy operations, 86 bacterial strains showing decarboxylase activity were isolated. From the wastewater samples, the species of genera Staphylococcus, Lactococcus, Enterococcus, Microbacterium, Kocuria, Acinetobacter, Pseudomonas, Aeromonas, Klebsiella and Enterobacter were identified by the MALDI-TOF MS and biochemical methods. The in vitro produced quantity of eight biogenic amines (BAs) was detected by the HPLC/UV-Vis method. All the isolated bacteria were able to produce four to eight BAs. Tyramine, putrescine and cadaverine belonged to the most frequently produced BAs. Of the isolated bacteria, 41% were able to produce BAs in amounts >100 mg L-1. Therefore, wastewater embodies a potential vector of transmission of decarboxylase positive microorganisms, which should be taken into consideration in hazard analyses within foodstuff safety control. The parameters of this wastewater (contents of nitrites, nitrates, phosphates, and proteins) were also monitored.
- MeSH
- Acinetobacter MeSH
- Aeromonas MeSH
- biogenní aminy chemie MeSH
- chemické látky znečišťující vodu chemie izolace a purifikace MeSH
- Enterobacter MeSH
- Enterococcus MeSH
- karboxylyasy chemie MeSH
- Klebsiella MeSH
- Lactobacillus MeSH
- Lactococcus MeSH
- Microbacterium MeSH
- mikrobiologie vody MeSH
- mlékárenství * MeSH
- odpadní voda analýza mikrobiologie MeSH
- Pediococcus MeSH
- Pseudomonas MeSH
- spektrofotometrie ultrafialová MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- Staphylococcus MeSH
- Streptococcus MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Evropa MeSH
Oxacillinases (OXA) have been mostly described in Enterobacteriaceae, Acinetobacter, and Pseudomonas species. Recent years have witnessed an increased prevalence of intrinsic and/or acquired β-lactamase-producing Acinetobacter in food-producing animals. This study was conducted to assess the prevalence of OXA among selected bacterial species and to characterize these enzymes by in silico analysis. Screening of OXA was performed by PCR amplification using specific pairs of oligonucleotides. Overall, 40 pairs of primers were designed, of which 6 were experimentally tested in vitro. Among 49 bacterial isolates examined, the presence of blaOXA-1-like genes was confirmed in 20 cases (41%; 19 times in Klebsiella pneumoniae and once in Enterobacter cloacae). No OXA were found in animal isolates. The study results confirmed the specificity of the designed oligonucleotide pairs. Furthermore, the designed primers were found to possess the ability to specifically detect 90.2% of all OXA. These facts suggest that the in silico and in vitro tested primers could be used for single or multiplex PCR to screen for the presence of OXA in various bacteria, as well as to monitor their spread. At the same time, the presence of conserved characteristic amino acids and motifs was confirmed by in silico analysis of sequences of representative members of OXA.
- MeSH
- antibakteriální látky farmakologie MeSH
- bakteriální proteiny genetika metabolismus MeSH
- beta-laktamasy genetika metabolismus MeSH
- DNA primery chemická syntéza metabolismus MeSH
- Enterobacter cloacae klasifikace účinky léků enzymologie genetika MeSH
- Escherichia coli klasifikace účinky léků enzymologie genetika MeSH
- exprese genu MeSH
- fylogeneze MeSH
- gramnegativní bakteriální infekce diagnóza epidemiologie mikrobiologie veterinární MeSH
- Klebsiella pneumoniae klasifikace účinky léků enzymologie genetika MeSH
- kur domácí mikrobiologie MeSH
- lidé MeSH
- maso mikrobiologie MeSH
- mikrobiální testy citlivosti MeSH
- multiplexová polymerázová řetězová reakce metody MeSH
- peniciliny farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
