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PubMed1977

[In-vitro sensitivity of Klebsiella-Enterobacter strains against cefazolin (author's transl)].

Investigation of the resistance of 290 strains of the Klebsiella-Enterobacter group (252 Klebsiella and 38 Enterobacter strains) against cefazolin showed that 63% of the Enterobacter strains were resistant and 50% of the Klebsiella strains were sensitive both in the serial dilution test and in the agar diffusion test. A total of 78% were inhibited by 32 mug cefazolin per millilitre. Isolates from the genitourinary tract were significantly more resistant than those from the respiratory tract.

Cefazolin↗

PubMed1998

Mechanisms of quinolone resistance in clinical isolates of Enterobacter cloacae.

OBJECTIVE: To study and evaluate changes in the gyrA gene and the outer-membrane protein patterns in relation to evolution of resistance against the quinolones in Enterobacter cloacae. METHODS: Strains expressing gyrA-mediated quinolone resistance become susceptible to quinolones upon insertion of the plasmid pNJR3-2. This plasmid (containing wild-type Escherichia coli quinolone-susceptible DNA gyrase A subunits) and pLA2917 (the vector) were introduced into 10 resistant or moderately susceptible clinical isolates of Enterobacter cloacae by conjugation. The transconjugants, the original isolates, the plasmid and the vector control were screened for susceptibility to ofloxacin, ciprofloxacin and sparfloxacin. Additionally, examinations of the outer-membrane proteins were performed. RESULTS: A reduction of MICs by a factor of 8--32 was found for the transconjugants of five Enterobacter cloacae isolates in the presence of the gene probe, suggesting that these isolates harbored mutations in gyrA. No discernible difference in the patterns of outer-membrane proteins of sensitive and resistant strains could be detected. CONCLUSIONS: It seems that changes in the target site such as alterations in gyrA are important factors leading to a change in the susceptibility of bacteria to the quinolones, whereas there were no evident changes in the outer-membrane proteins to account for evolution of resistance.

Journal Article↗

PubMed1976

Susceptibility of Enterobacter to cefamandole: evidence for a high mutation rate to resistance.

Cefamandole minimum inhibitory concentrations (MICs) of 10 strains of Enterobacter were determined by the ICS agar dilution and broth dilution procedures. Agar dilution MICs ranged from 1 to 8 mug/ml, with an inoculum of 10(4) organisms/spot. Broth dilution MICs were consistently higher, with an inoculum of approximately 7 x 10(5) organisms/ml. Seven strains showed MICs of >/=64 mug/ml. There was a marked inoculum effect in broth, and skipped tubes were often observed. Variants resistant to 32 mug/ml or more were isolated by direct selection and were shown to occur at a frequency of approximately 10(-6) to 10(-7). A mutant showing a 16-fold increase in agar dilution MIC was also isolated by indirect selection. These variants and others isolated from broth in the presence of cefamandole were tested for ability to inactivate the antibiotic, using both a biological and a chemical procedure. Two distinct classes of variants were seen. Twelve of 28 were shown by both methods to inactivate the antibiotic, whereas the others, including the indirectly selected mutant, did not. The wild types were also negative by both tests. The higher cefamandole MICs of Enterobacter in broth, thus, appeared to reflect a high frequency of resistant variants that were not detected with the inoculum and end point criteria usually used in agar dilution methods. The ability of some variants to inactivate cefamandole may have resulted from a mutation that extended the activity of Enterobacter cephalosporinase to include this antibiotic.

Cephalosporins↗

PubMed1970

Relationship Between beta-Lactamase Activity and Resistance of Enterobacter to Cephalothin.

The relationship between cephalosporin beta-lactamase activity and resistance to cephalothin was investigated in strains of Enterobacter cloacae and E. aerogenes. beta-Lactamase activity was detected in all strains, but a quantitative correlation between amount of beta-lactamase activity and level of resistance to cephalothin was not observed. Permeability barriers to cephalothin were observed and varied from strain to strain. beta-Lactamase activity was increased by growing organisms in the presence of penicillin G. These enzymes hydrolyzed cephalosporins more rapidly than penicillins. Penicillinase-resistant penicillins, especially those of the isoxazolyl series, effectively inhibited Enterobacter beta-lactamase. A synergistic antibacterial effect was observed when organisms were exposed to cephalothin and oxacillin in combination, and the resistance of even very small inocula to cephalothin was reduced by addition of oxacillin. Oxacillin probably exerts its effect by inhibiting beta-lactamase at an intracellular site. Intracellular beta-lactamase may make an important contribution to the resistance of even small inocula of gram-negative bacilli to cephalosporin and penicillin antibiotics. Although beta-lactamase plays a significant role in the resistance of Enterobacter to cephalothin, other factors, such as permeability barriers, also participate in determining the level of resistance.

Journal Article↗

PubMed1996

Outbreak of TEM-24-producing Enterobacter aerogenes in an intensive care unit and dissemination of the extended-spectrum beta-lactamase to other members of the family enterobacteriaceae.

We report an outbreak of Enterobacter aerogenes in an intensive care unit (ICU) and two medicine departments that produced the extended-spectrum beta-lactamase TEM-24, which was difficult to detect by disk agar diffusion. The strains were compared by DNA restriction fragment length polymorphism after pulsed-field gel electrophoresis following cleavage with XbaI. This typing method indicated that a single strain, first isolated in the ICU, spread throughout the other medical departments as a result of patient transfer. We also observed the transfer in vivo of the plasmid encoding TEM-24 from the strain of Enterobacter aerogenes to different strains of Escherichia coli and Citrobacter freundii in the ICU. It therefore appears that the epidemic involved results from two events: dissemination of one strain of Enterobacter aerogenes and dissemination of the plasmid encoding TEM-24 among various members of the family Enterobacteriaceae.

Anti-Bacterial Agents↗

PubMed1996

Adhesive properties and antibiotic resistance of Klebsiella, Enterobacter, and Serratia clinical isolates involved in nosocomial infections.

Intestinal colonization by Klebsiella, Enterobacter, and Serratia (KES) strains is a crucial step in the development of nosocomial infections. We studied the adhesive properties, antibiotic resistance, and involvement in colonization or infection of 103 KES clinical isolates: 30 Klebsiella pneumoniae (29%), 16 Klebsiella oxytoca (15%), 30 Enterobacter aerogenes (29%), 14 Enterobacter cloacae (14%), and 13 Serratia sp. (13%) isolates. Half of them were resistant to several antimicrobial agents, including aminoglycosides and beta-lactam antibiotics. A total of 27 of 30 K. pneumoniae isolates (90%) adhered to the human cell line Intestine-407 (Int-407), while none of the K. oxytoca or E. aerogenes isolates and only 2 of the E. cloacae isolates adhered. Three adhesive patterns were observed for K. pneumoniae: an aggregative adhesion in 57% of the isolates, a diffuse adhesion in only one isolate, and a new pattern, localized adhesion, in 30% of the isolates. While most of the sensitive strains adhered with the aggregative phenotype, the localized pattern was associated with resistant K. pneumoniae isolates producing the CAZ-5 beta-lactamase. Furthermore, 45% of such localized-adhesion isolates were involved in severe infections. The distributions of type 1 and type 3 fimbriae, enteroaggregative E. coli, and cf29, pap, and afa/Dr adhesin-encoding genes were determined by using specific DNA probes. No relationship was found between the adhesive pattern and the production of specific fimbriae, suggesting that several unrecognized adhesive factors are involved. Our study indicates that special adhesive properties associated with resistance to antimicrobial agents could account for the pathogenicity of certain nosocomial strains.

Adhesins, Bacterial↗

PubMed1997

Phylogenetical relationship based on groE genes among phenotypically related Enterobacter, Pantoea, Klebsiella, Serratia and Erwinia species.

In an attempt to define the phylogenetical relationship among 17 phenotypically related species of genera Enterobacter, Pantoea, Serratia, Klebsiella and Erwinia, we determined almost all of their groE operon sequences using the polymerase chain reaction direct sequencing method. The number of nucleotide substitutions per site was 0.12+/-0.030. The value was 3.6-fold higher than that of 16S rDNA. As a result, we were successful in constructing molecular phylogenetic trees which had a finer resolution than that based on the 16S rDNA sequences. The phylogenetic trees based on the nucleotide sequences and deduced amino acid sequences of groE operons indicated that the members of genera Enterobacter, Pantoea and Klebsiella were closely related to each other, while Serratia and Erwinia species except Erwinia carotovora, made distinct clades. The close relationship between Enterobacter aerogenes and Klebsiella pneumoniae, that had been suggested by biochemical tests and DNA hybridization, was also supported by our molecular phylogenetic trees.

Journal Article↗

PubMed1985

[Enterobacter cloacae and E. aerogenes septicemia: emergence of resistant variants (derepressed cephalosporinase) during treatment with third-generation cephalosporins].

From three patients hospitalised in intensive care units with Enterobacter septicaemia (two cases with E. cloacae, and one with E. aerogenes), cefotaxime therapy, alone or in combination with an aminoglycoside, selected variants (R) with increased resistance to beta-lactam antibiotics. The cross-resistance extended to all the beta-lactam antibiotics tested, penicillins and cephalosporins, including third-generation cephalosporins. The crude extracts of uninduced cultures of R variants showed high beta-lactamase activity and of the cephalosporinase type. These variants were selected in vitro with a frequency of 10(-6) to 10(-7) and may result from a mutation involving the regulation of Enterobacter cephalosporinases, usually inducible. Data from the literature indicated that this new type of resistance is actually emerging and observed not only in Enterobacter sp. The problem of emergence of R variants exhibiting cross-resistance to beta-lactam antibiotics should be considered when third-generation cephalosporins are used.

Adult↗

PubMed2003

Factors influencing gene expression and resistance for Gram-negative organisms expressing plasmid-encoded ampC genes of Enterobacter origin.

High-level expression of AmpC beta-lactamases results in organisms resistant to multiple beta-lactam antibiotics. The mechanism of chromosomally mediated AmpC resistance has been elucidated, however the mechanism(s) driving plasmid-encoded AmpC resistance are unknown. Studies were designed to identify factors which influence expression of plasmid-encoded ampC genes and correlate these factors with resistance. As the model system, ampC genes of Enterobacter origin were used to determine how gene copy number, genetic background and genetic organization influenced resistance phenotypes. To this end, gene expression from the plasmid-encoded inducible blaACT-1 and non-inducible blaMIR-1 were compared with chromosomal ampC gene expression from both wild-type (WT) and derepressed Enterobacter cloacae isolates. RNA levels within the original clinical isolates were examined using primer extension analysis, whereas a new PCR strategy was developed to examine gene copy number. These data revealed that blaACT-1 and blaMIR-1 constitutive expression was 33- and 95-fold higher than WT expression, whereas copy numbers of the plasmid-encoded genes were 2 and 12, respectively. Differences in promoters and transcriptional starts for the respective plasmid-encoded genes were noted and contribute to increases observed in overall expression. Finally, beta-lactam MICs were increased two- to 16-fold when blaACT-1 was expressed in Escherichia coli AmpD- strains compared with E. coli AmpD+ strains. In conclusion, high-level expression of plasmid-encoded ampC genes requires interplay between multiple factors including genetic organization, promoter modifications, genetic background, and to some extent gene copy number. In addition, clinical laboratories need to be aware that genetic backgrounds of inducible plasmid-encoded genes can dramatically influence MICs for organisms not normally associated with derepressed phenotypes.

Anti-Bacterial Agents↗

PubMed2004

Incidence of antibiotic-resistant Klebsiella pneumoniae and Enterobacter species in freshwater wetlands.

AIMS: The aim of this study was to assess the incidence of Enterobacteriaceae (potential human and animal pathogens) in wetlands. METHODS: Enterobacteriaceae, selected from the sediments and rhizosphere of wetland plant Juncus effusus L., were analysed using classical microbiological methods, API20E, API20NE, fatty acid analyses, and 16S rRNA sequencing. Assessed virulence factors include antibiotic resistance, presence of plasmids and capsules. RESULTS: Klebsiella pneumoniae, Enterobacter cloacae and Enterobacter asburiae, known human pathogens, were identified. K. pneumoniae 16S rRNA gene sequence showed the significant hit (E < 0.001) with the unculturable bacteria obtained from faeces of elderly individuals (accession number AB099804) when Genbank database was used. Ent. asburiae 16S rRNA gene sequence showed the significant hit with (E < 0.001) with the unculturable bacteria obtained from the pig gastrointestinal tract (accession number AF371852). The rate of antibiotic resistance (<50 microg ml(-1)) was high for ampicillin and cephalosporins for the most strains (75.7%) yet low (>10 to 20 microg ml(-1)) for kanamycin, tetracycline and chloramphenicol for all strains tested. Capsules were detected in all investigated strains. PCR detected membrane protein but not chromosomally encoded beta-lactamase. SIGNIFICANCE AND IMPACT OF THE STUDY: The antibiotic resistance of tested strains and presence of capsules (protect micro-organisms from phagocytosis) suggest that wetland sediments and rhizosphere present a potential reservoirs for enteric human and animal pathogens.

Bacterial Capsules↗

PubMed2025

Isolation and characterization of strictly lytic bacteriophages against carbapenem-resistant Enterobacter cloacae complex.

UNLABELLED: The global surge of carbapenem-resistant Enterobacter cloacae complex (CR-ECC) poses a significant clinical challenge due to limited treatment options. This study aimed to isolate and characterize lytic bacteriophages (phages) targeting CR-ECC. CR-ECC CYEBC080 was used as the bacterial host for isolating lytic phages, and a comprehensive evaluation was conducted on isolated phages, including phage stability under various pH and temperature conditions, host range analysis, killing curves, and therapeutic efficacy in Galleria mellonella larvae and a murine bacteremia model. Twelve lytic phages with distinct random amplified polymorphic DNA patterns were isolated, and transmission electron microscopy confirmed their classification under the Straboviridae family within the Caudoviricetes class. All phages remained stable across pH 3-11 for up to 90 minutes, with an optimal temperature range of 25°C-37°C. Among them, CYPEBC012 exhibited the broadest host range, lysing 93.75% of 80 CR-ECC isolates, while CYPEBC006 displayed the narrowest, lysing only 65%. Whole-genome sequencing revealed 12 phages with linear double-stranded DNA genomes ranging from 177,624 to 180,648 bp. Phage treatment administered at a multiplicity of infection of 10, 1 hour post-infection, significantly improved larval survival at day 7, reaching ≥80% in most groups, except CYPEBC001 (50%) and CYPEBC004 (60%) treatment groups. In CYEBC080-infected mice, CYPEBC012 treatment resulted in 100% survival by day 3 and 80% survival through day 7. Additionally, phage-treated mice exhibited significantly reduced bacterial loads and high phage titers in blood and liver. This study demonstrates the therapeutic potential of CYPEBC012 as a promising strategy against CR-ECC infections, offering an alternative to conventional antimicrobial treatments. IMPORTANCE: This study identified and characterized lytic bacteriophages targeting carbapenem-resistant Enterobacter cloacae complex, with CYPEBC012 exhibiting the broadest host range and significantly improving survival in a murine bacteremia model. Its stability and efficacy highlight its potential for clinical application. Our findings demonstrate that phage therapy offers a promising alternative to conventional treatments to combat antibiotic-resistant infections.

Animals↗

PubMed1992

Beta-lactamase expression and cross-resistance to beta-lactam antibiotics in a nosocomial population of Enterobacter.

This study sought to quantitatively describe the spectrum of constitutive and inducible beta-lactamase activity present in a tertiary care center's population of Enterobacter species. beta-Lactamase activity in the absence and presence of 2 recognized beta-lactamase-inducing antibiotics, cefoxitin and imipenem, was measured. The Enterobacter cloacae (n = 35) population was strikingly bimodal, expressing 'all-or-none' cross-resistance to beta-lactams (except imipenem) corresponding to the baseline level of beta-lactamase expression. E. aerogenes (n = 14) displayed a less dichotomous pattern of resistance, and MICs of beta-lactam antibiotics were less strongly related to the magnitude of enzyme activity. We conclude that our nosocomial population of E. cloacae, like strains rendered resistant to beta-lactams in vitro, is largely dependent upon beta-lactamase as a mechanism of this resistance. Furthermore, we document the presence of a large subpopulation of beta-lactam-susceptible isolates possessing inducible beta-lactamase and therefore subject to selection for enzyme derepression.

Anti-Bacterial Agents↗

PubMed2003

[Evaluation of relations between plasmids and phage host range among clinical isolates of Enterobacter cloacae].

The aim of this study was evaluation the plasmid influence on phage host range of clinical strains of Enterobacter cloacae. We found that strains included in restrictive pattern A, displayed reduced host range. Such reduced sensitivity make these strains excellent candidates for search restrictive-modification systems. High discriminative efficacy of isolated phages (specific for strains Enterobacter cloacae) make them useful tool for phage typing in epidemiological investigations.

Bacteriophage Typing↗

PubMed1996

Acid and base hydrolysis of lipid A from Enterobacter agglomerans as monitored by electrospray ionization mass spectrometry: pertinence to detoxification mechanisms.

Lipopolysaccharides (LPS), which are endotoxins found in the cell wall of Gram-negative bacteria, are common components of organic dusts that cause or contribute to symptoms associated with organic dust diseases. The lipid A subgroup within LPS is believed to be responsible for the toxicity. Acid and base treatments, which can be effective detoxification methods, were performed on lipid A from Enterobacter agglomerans (EA), a bacterium commonly found in field cotton. Negative-ion electrospray ionization mass spectrometry was employed to characterize the post-treatment structural changes to lipid A. Acid treatment (1% acetic acid, 100 degrees C) hydrolyzed the ester side-chains of lipid A. It was found that the ester-linked palmitoyl group was the most labile to acid hydrolysis. Hydrolysis of the palmitoyl moiety conformed to pseudo-first-order chemical reaction kinetics with a rate constant for decomposition of heptacyl-lipid A from Enterobacter agglomerans of approximately 3.3 x 10(-3) min-1. An order of lability of lipid A acyl side-chains to acid hydrolysis was also deduced: R4' (palmitoyl) > R1' (myristoyl or hydroxymyristoyl) > R3 (hydroxymyristoyl at position 3) > R1 (oxymyristoyl group at position 3') > R2' (lauroyl). Base treatment (0.05 M NaOH in 95% EtOH, 65 degrees C) was shown to be more effective at cleaving ester-linked side-chains. In addition, mass spectral evidence suggests that opening of the pyranose rings of the disaccharide backbone of lipid A and/or removal of the phosphoryl groups may be occurring during base treatment. This study sheds light on mechanistic aspects of treatment procedures leading to the detoxification of endotoxins.

Acids↗

PubMed2026

Genomic and Phenotypic Characterization of Two Novel Enterobacter Phages With EDTA-Enhanced Antibiofilm Activity.

Multidrug-resistant members of the Enterobacter cloacae complex (ECC) are increasingly linked to difficult-to-treat infections and biofilm-mediated antimicrobial tolerance. Here, two lytic phages, vB_EhoIP_HHH and vB_EluM_RZH, displaying podovirus-like and myovirus-like morphology, respectively, were isolated from the River Chelt. HHH has a 39,582 bp genome (51.2% GC, 63 ORFs), while RZH has a 174,197 bp genome (39.4% GC, 314 ORFs), with neither genome carrying antimicrobial resistance, virulence or lysogeny-associated genes. VIRIDIC and VICTOR analyses placed HHH within Kayfunavirus and RZH within Karamvirus, supporting their classification as distinct species. Both phages demonstrated rapid adsorption, short latent periods and stability across physiological pH and temperature ranges. A phage cocktail targeting MDR ECC strain was evaluated with EDTA against established biofilms. Crystal violet assays showed the greatest biomass reduction at MOI 10 with 0.5-0.75 mM EDTA. Bliss independence analysis revealed localized synergy within this window but significant overall antagonism at higher EDTA concentrations. CFU enumeration confirmed greater activity against 24 h than 48 h biofilms. The optimized combination also reduced recoverable bacteria in a fibroblast infection model while maintaining low LDH release. These findings identify two novel lytic Enterobacter phages and support a narrow EDTA concentration window for enhanced phage-mediated antibiofilm activity.

Biofilms↗

PubMed2004

Effect of high concentration of Co (II) on Enterobacter liquefaciens strain C-1: a bacterium highly resistant to heavy metals with an unknown genome.

Heavy metals are required as nutrients for essential functions in microorganisms. However, higher concentrations of these cations are generally toxic and may produce contrasting effects on living organisms. Enterobacter liquefaciens strain C-1, a bacterium isolated from the Moa mine in Cuba, is able to survive in the presence of high concentrations of heavy metals. The proteomes of Enterobacter liquefaciens strain C-1, grown under aerobic conditions in the presence and absence of Co (II) were compared using two-dimensional gel electrophoresis analysis in the isoelectric point range of 4-7 and the mass range of 15-120 kDa. Significant changes in the expression level (> two-fold) were detected for 13 spots: seven and six were up- and down-regulated, respectively. Because the genome of this bacterium is unknown, identification by peptide mass fingerprinting only succeeded in four cases and most of the cross-species identifications were supported by de novo sequencing of tryptic peptides followed by sequence alignment using the MS BLAST program. Twelve different proteins were identified, ten are involved in cellular antioxidant defence probably induced by the presence of Co (II). This is the first step towards understanding the role of proteins participating in the mechanism of resistance to heavy metals in this bacterium.

Amino Acid Sequence↗

PubMed1984

Biosynthesis of bacterial glycogen: characterization of adenosine diphosphate glucose synthetases from Enterobacter hafniae and Aeromonas hydrophila.

Enterobacter hafniae and Aeromonas hydrophila ADPglucose synthetases were purified approximately 39- and 61-fold, respectively, over the crude extract. Both enzymes were heat stable at 60 degrees C in the presence of inorganic phosphate. The molecular weights of both enzymes were approximately 200,000 which are similar to other enteric ADPglucose synthetases studied. Based on kinetic results obtained from the partially purified enzymes, the E. hafniae enzyme is activated twofold by phospho-enolpyruvate while the A. hydrophila enzyme is activated twofold by fructose 6-P and 1.5-fold by fructose 1,6 bis-phosphate. The E. hafniae enzyme activity is strongly inhibited by AMP and ADP and the inhibition can be partially reversed by P-enolpyruvate. ADP is the most effective inhibitor of the A. hydrophila enzyme and its inhibition can be partially overcome by the presence of the activators fructose 6-P and fructose 1,6-P2. These kinetic results show that the allosteric properties of the E. hafniae enzyme are distinctly different from the ADPglucose synthetases of those previously studied from bacteria of the genus Enterobacter. Although the A. hydrophila enzyme is activated by fructose 1,6-P2, its allosteric properties are quite different than those observed for ADPglucose synthetase of the Enterobacteriaceae.

Adenosine Diphosphate↗

PubMed1988

Serotyping and phage typing to identify Enterobacter cloacae contaminating total parenteral nutrition.

The origin of an outbreak of Enterobacter cloacae septicemia in six surgery patients was traced down to the total parenteral nutrition production line. While the endproduct of the production line was sterile, Enterobacter cloacae and other bacteria were detected on different tabs and tubings of the line. It is believed that the bacteria were transferred from the tabs to a few bags of the total parenteral nutrition in one batch by touch contamination. Serotyping and phage-typing of the clinical isolates revealed that five of the patients' strains were identical.

Bacteriophage Typing↗