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
- Bacteriophages * genetics MeSH
- Enterobacter cloacae MeSH
- Yogurt MeSH
- Milk microbiology MeSH
- Refuse Disposal * MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article 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
- Drug Resistance, Bacterial MeSH
- Enterobacter cloacae * drug effects MeSH
- Antimicrobial Cationic Peptides pharmacology therapeutic use MeSH
- Drug Therapy, Combination MeSH
- Humans MeSH
- Staphylococcus aureus * drug effects MeSH
- Drug Synergism * MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't 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
- Anti-Bacterial Agents pharmacology MeSH
- Bacterial Proteins genetics metabolism MeSH
- beta-Lactamases genetics metabolism MeSH
- DNA Primers chemical synthesis metabolism MeSH
- Enterobacter cloacae classification drug effects enzymology genetics MeSH
- Escherichia coli classification drug effects enzymology genetics MeSH
- Gene Expression MeSH
- Phylogeny MeSH
- Gram-Negative Bacterial Infections diagnosis epidemiology microbiology veterinary MeSH
- Klebsiella pneumoniae classification drug effects enzymology genetics MeSH
- Chickens microbiology MeSH
- Humans MeSH
- Meat microbiology MeSH
- Microbial Sensitivity Tests MeSH
- Multiplex Polymerase Chain Reaction methods MeSH
- Penicillins pharmacology MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Czech Republic MeSH
Biofilm is involved in a variety of infections, playing a critical role in the chronicity of infections. Enterobacter cloacae is a biofilm-forming and multi-drug-resistant (MDR) nosocomial pathogen leading to significant morbidity and mortality. This study aimed at isolation of a bacteriophage against MDR clinical strain of E. cloacae and its efficacy against bacterial planktonic cells and biofilm. A bacteriophage MJ2 was successfully isolated from wastewater and was characterized. The phage exhibited a wide range of thermal and pH stability and demonstrated considerable adsorption to host bacteria in the presence of CaCl2 or MgCl2. Transmission electron microscopy (TEM) showed MJ2 head as approximately 62 and 54 nm width and length, respectively. It had a short non-contractile tail and was characterized as a member of the family Podoviridae [order Caudovirales]. The phage MJ2 was found to possess 11 structural proteins (12-150 kDa) and a double-stranded DNA genome with an approximate size of 40 kb. The log-phase growth of E. cloacae both in biofilm and suspension was significantly reduced by the phage. The E. cloacae biofilm was formed under different conditions to evaluate the efficacy of MJ2 phage. Variable reduction pattern of E. cloacae biofilm was observed while treating it for 4 h with MJ2, i.e., biofilm under static conditions. The renewed media with intervals of 24, 72, and 120 h showed biomass decline of 2.8-, 3-, and 3.5-log, respectively. Whereas, the bacterial biofilm formed with dynamic conditions with refreshing media after 24, 72, and 120 h demonstrated decline in growth at 2.5-, 2.6-, and 3.3-log, respectively. It was, therefore, concluded that phage MJ2 possessed considerable inhibitory effects on MDR E. cloacae both in planktonic and biofilm forms.
- MeSH
- Biofilms growth & development MeSH
- Magnesium Chloride pharmacology MeSH
- Calcium Chloride pharmacology MeSH
- Genome Size MeSH
- DNA, Viral MeSH
- Enterobacter cloacae virology MeSH
- Genome, Viral genetics MeSH
- Host Specificity MeSH
- Hydrogen-Ion Concentration MeSH
- Microbial Viability MeSH
- Drug Resistance, Multiple, Bacterial * MeSH
- Molecular Weight MeSH
- Wastewater virology MeSH
- Podoviridae isolation & purification physiology ultrastructure MeSH
- Virus Attachment drug effects MeSH
- Temperature MeSH
- Viral Proteins chemistry MeSH
- Publication type
- Journal Article MeSH
ST252 Enterobacter cloacae, producing GES-5 carbapenemase, was isolated in a Czech hospital. blaGES-5was part of a novel class 1 integron, In1406, which also included a new allele of the aadA15 gene cassette. In1406 was located on a ColE2-like plasmid, pEcl-35771cz (6953bp).
- MeSH
- Bacterial Proteins genetics MeSH
- beta-Lactamases genetics MeSH
- Enterobacter cloacae enzymology genetics MeSH
- Enterobacteriaceae Infections microbiology MeSH
- Integrons genetics MeSH
- Humans MeSH
- Multilocus Sequence Typing MeSH
- Hospitals MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Czech Republic MeSH
A VIM-1-producing ST92 Enterobacter cloacae was isolated in a Czech hospital. blaVIM-1 was part of the class 1 integron In110 carried by a Tn1721-like transposon. Tn1721-like was located on a ColE1-like plasmid, pEncl-30969cz (33,003 bp). Target site duplications at the boundaries of Tn1721-like suggested its transposition into the pEncl-30969cz backbone.
- MeSH
- Anti-Bacterial Agents pharmacology MeSH
- Bacterial Proteins genetics MeSH
- beta-Lactamases genetics MeSH
- beta-Lactam Resistance genetics MeSH
- DNA, Bacterial analysis genetics MeSH
- Enterobacter cloacae genetics MeSH
- Enterobacteriaceae Infections microbiology MeSH
- Integrons genetics MeSH
- Humans MeSH
- Microbial Sensitivity Tests MeSH
- Plasmids genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
The imbalance of the intestinal microbiota is link, by several diseases such as celiac disease, which causes inflammation in the small intestine. This inflammation is due by the digestion of gluten present in some type of cereals. In this work, fecal and duodenal biopsy samples were collected to characterize by conventional culture technique the composition of the Enterobacteries group in intestinal mirobiota of celiac children and were compared with control children. A significant difference detected in the intestinal flora of celiac children compared to controls children concerning the Enterobacteries group. We found an increase of E.coli, Enterobacter aerogeneses, and Klebsiella with presence of Salmonella sp, Shigella sp in biopsy and fecal samples of celiac children and a relationship between the increase of Enterobacter cloacea and the presence of positive anti-transglutaminase value.
- MeSH
- Bacteriological Techniques * MeSH
- Celiac Disease * pathology MeSH
- Citrobacter isolation & purification MeSH
- Child MeSH
- Duodenum microbiology MeSH
- Enterobacter aerogenes isolation & purification MeSH
- Enterobacter cloacae isolation & purification MeSH
- Escherichia coli isolation & purification MeSH
- Feces microbiology MeSH
- Klebsiella isolation & purification MeSH
- Humans MeSH
- Salmonella isolation & purification MeSH
- Shigella isolation & purification MeSH
- Gastrointestinal Microbiome * MeSH
- Intestinal Mucosa microbiology MeSH
- Case-Control Studies MeSH
- Transglutaminases MeSH
- Check Tag
- Child MeSH
- Humans MeSH
The complete nucleotide sequences of three multidrug resistance (MDR) IncA/C-like plasmids from Enterobacteriaceae isolates carrying the VIM-type carbapenemase-encoding integrons In4863 (blaVIM-19-aacA7-dfrA1-ΔaadA1-smr2) or In4873 (blaVIM-1-aacA7-dfrA1-ΔaadA1-smr2) were determined, which are the first In416-like elements identified in Greece. Plasmids pKP-Gr642 and pKP-Gr8143 were from Klebsiella pneumoniae ST383 isolates, whereas plasmid pEcl-Gr4873 was from an Enterobacter cloacae ST88 isolate. Sequencing showed that pKP-Gr642 (162787bp) and pKP-Gr8143 (154395bp) consisted of the type 1 IncA/C2 conserved backbone, the blaCMY-2-like gene-containing region, and the ARI-B (with the sul2 gene) and ARI-A (with a class 1 integron) resistance islands, like the plasmid pUMNK88_161 from the USA. The third plasmid, pEcl-Gr4873 (153958bp), exhibited extensive similarity with the type 2 IncA/C2 plasmid pR55 from France. pEcl-Gr4873 carried only one resistance island of a hybrid transposon structure inserted in a different location to ARI-A in type 1 A/C2 plasmids. In all three plasmids, the In416-like integrons In4863 or In4873 were identified within non-identical class II transposon structures. All three In416-like-carrying regions presented significant similarities with the MDR region of the IncA/C2 plasmid pCC416 from Italy, carrying the prototype In416 integron (blaVIM-4-aacA7-dfrA1-ΔaadA1-smr2). These findings provided the basis for speculations regarding the evolution of IncA/C2 plasmids with In416-like integrons, and confirmed the rapid evolution of some IncA/C2 plasmid lineages. Considering the broad host range of IncA/C2 molecules, it seems that pKP-Gr642, pKP-Gr8143 and pEcl-Gr4873 plasmids might support the diffusion of In416-like integrons among Enterobacteriaceae.
- MeSH
- beta-Lactamases genetics MeSH
- Enterobacter cloacae enzymology genetics isolation & purification MeSH
- Enterobacteriaceae Infections microbiology MeSH
- Integrons * MeSH
- Klebsiella pneumoniae enzymology genetics isolation & purification MeSH
- Humans MeSH
- Molecular Sequence Data MeSH
- Plasmids * MeSH
- Gene Order MeSH
- Sequence Analysis, DNA MeSH
- Sequence Homology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Greece MeSH
BACKGROUND: Increasing bacterial resistance to antibiotics is one of the most serious problems in current medicine. An important factor contributing to the growing prevalence of multiresistant bacteria is application of antibiotics. This study aimed at analyzing the development of resistance of Enterobacteriaceae to selected beta-lactam, fluoroquinolone and aminoglycoside antibiotics in the University Hospital Olomouc and assessing the effect of selection pressure of these antibiotics. METHODS: For the period between 1 January 2000 and 31 December 2011, resistance of Klebsiella pneumoniae, Escherichia coli, Enterobacter cloacae and Proteus mirabilis to third- and fourth-generation cephalosporins, meropenem, piperacillin/tazobactam, fluoroquinolones and aminoglycosides was retrospectively studied. For the assessment of selection pressure of antibiotics, a parameter of defined daily dose in absolute annual consumption (DDDatb) based on the ATC/DDD classification and in relative annual consumption (RDDDatb) as the number of defined daily doses per 100 bed-days was used. The relationship between frequency of strains resistant to a particular antibiotic and antibiotic consumption was assessed by linear regression analysis using Spearman's correlation. The level of statistical significance was set at p < 0.05. RESULTS: A total of 113,027 isolates from the Enterobacteriaceae family were analyzed. There was a significant effect of selection pressure of the primary antibiotic in the following cases: piperacillin/tazobactam in Klebsiella pneumoniae, gentamicin in Klebsiella pneumoniae and Escherichia coli and amikacin in Escherichia coli and Enterobacter cloacae. Also, there was significant correlation between resistance to ceftazidime and consumption of piperacillin/tazobactam in Klebsiella pneumoniae and Escherichia coli. No relationship was found between consumption of third- and fourth-generation cephalosporins and resistance to ceftazidime or between fluoroquinolone consumption and resistance to ciprofloxacin. CONCLUSION: The study showed the effects of both direct and indirect selection pressure on increasing resistance to gentamicin, amikacin, piperacillin/tazobactam and ceftazidime. Given the fact that no correlation was found between resistance to fluoroquinolones and consumption of either primary or secondary antibiotics, we assume that the increasing resistance to fluoroquinolones is probably due to circulation of resistance genes in the bacterial population and that this resistance was not affected by reduced use of these antibiotics.
- MeSH
- Aminoglycosides therapeutic use MeSH
- Anti-Bacterial Agents therapeutic use MeSH
- beta-Lactams therapeutic use MeSH
- Enterobacter cloacae drug effects isolation & purification physiology MeSH
- Enterobacteriaceae Infections drug therapy microbiology MeSH
- Escherichia coli drug effects isolation & purification physiology MeSH
- Fluoroquinolones therapeutic use MeSH
- Klebsiella pneumoniae drug effects isolation & purification physiology MeSH
- Humans MeSH
- Linear Models MeSH
- Microbial Sensitivity Tests MeSH
- Drug Resistance, Multiple, Bacterial drug effects MeSH
- Proteus mirabilis drug effects isolation & purification physiology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
