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PubMed1998

Detection of intimins alpha, beta, gamma, and delta, four intimin derivatives expressed by attaching and effacing microbial pathogens.

Intimins are outer membrane proteins expressed by enteric bacterial pathogens capable of inducing intestinal attachment-and-effacement lesions. A eukaryotic cell-binding domain is located within a 280-amino-acid (Int280) carboxy terminus of intimin polypeptides. Polyclonal antiserum was raised against Int280 from enteropathogenic Escherichia coli (EPEC) serotypes O127:H6 and O114:H2 (anti-Int280-H6 and anti-Int280-H2, respectively), and Western blot analysis was used to explore the immunological relationship between the intimin polypeptides expressed by different clinical EPEC and enterohemorrhagic E. coli (EHEC) isolates, a rabbit diarrheagenic E. coli strain (RDEC-1), and Citrobacter rodentium. Anti-Int280-H6 serum reacted strongly with some EPEC serotypes, whereas anti-Int280-H2 serum reacted strongly with strains belonging to different EPEC and EHEC serotypes, RDEC-1, and C. rodentium. These observations were confirmed by using purified Int280 in an enzyme-linked immunosorbent assay and by immunogold and immunofluorescence labelling of whole bacterial cells. Some bacterial strains were recognized poorly by either antiserum (e.g., EPEC O86:H34 and EHEC O157:H7). By using PCR primers designed on the basis of the intimin-encoding eae gene sequences of serotype O127:H6, O114:H2, and O86:H34 EPEC and serotype O157:H7 EHEC, we could distinguish between different eae gene derivatives. Accordingly, the different intimin types were designated alpha, beta, delta, and gamma, respectively.

Adhesins, Bacterial↗
PubMed2005

Dissemination of CTX-M-type extended-spectrum beta-lactamase genes to unusual hosts.

A Citrobacter amalonaticus and a Morganella morganii producing the CTX-M-1 extended-spectrum beta-lactamase (ESBL) were isolated from an area where this enzyme is now widespread in Escherichia coli. This is the first report of CTX-M-1 in the former species. In both cases the ESBL determinant was possibly acquired by these unusual hosts in vivo, after coinfection with E. coli strains carrying conjugative plasmids encoding CTX-M-1.

Citrobacter↗
PubMed1977

Effect of storage conditions of the performance of bismuth sulfite agar.

Refrigerated storage of bismuth sulfite agar plates for up to 4 days did not adversely affect growth and colonial characteristics of selected Salmonella strains. Incubation of inoculated plates for 48 h favored the development of more salmonellae with typical morphology. Inoculated plates of freshly poured medium incubated for 48 h gave recoveries similar to those on refrigerated plates and showed a high selectivity against Citrobacter freundii and Proteus vulgaris, organisms which mimic the colonial characteristics of Salmonella on this medium. The use of bismuth sulfite plates stored at room temperature for more than 2 days should be avoided.

Agar↗
PubMed1977

Esculin hydrolysis by Enterobacteriaceae.

Literature reports disagree concerning esculin hydrolysis in the family Enterobacteriaceae. A total of 2,490 strains of the family were investigated for esculin hydrolysis by two methods, the esculin spot test and the PathoTec incubation strip, which measures constitutive enzyme, and five growth-supporting methods, which determine both constitutive and inducible enzymes. The five growth-supporting media studied were: Vaughn-Levine, the standard esculin hydrolysis medium (P. R. Edwards and W. H. Ewing, Identification of Enterobacteriaceae, 3rd ed., 1972); Vaughn-Levine without iron; Vaughn-Levine without Andrade's indicator; and bile-esculin medium. Growth media were incubated at 35 degrees C and checked every 24 h for 120 h. On growth media, 0.3% of Escherichia coli were positive in 24 h, 34% in 48 h, and 61% in 120 h. No strains were positive on the "nongrowth" tests. It appeared that the esculin hydrolysis enzyme(s) of E. coli was inducible rather than constitutive. All esculin hydrolyzers, which yielded positive tests on "constitutive tests" and 24-h tests, were limited to the genera Klebsiella, Enterobacter, and Serratia and species of Proteus vulgaris, Proteus rettgeri, and Citrobacter diversus. When used with standardized inoculum size and incubation time, the esculin hydrolysis test is very useful for differentiation within the family Enterobacteriaceae.

Citrobacter↗
PubMed2005

Detection and identification of bacterial DNA in serum from patients with acute pancreatitis.

BACKGROUND AND AIMS: Bacterial infections are common complications in patients with acute pancreatitis, and translocation of bacteria from the intestinal lumen is probably the first step in the pathogenesis of these infections. As blood cultures in afebrile patients are usually negative, more sensitive methods to investigate this hypothesis in patients are needed. Our group has recently developed a method to detect the presence of bacterial DNA in biological fluids, and we aimed to detect bacterial DNA in patients with acute pancreatitis, as molecular evidences of bacterial translocation. METHODS: Samples of blood were obtained on three consecutive days within the first six days after admission. Bacterial DNA was detected using a polymerase chain reaction based method, and an automated DNA nucleotide sequencing process allowed identification of bacteria species. RESULTS: Thirty one consecutively admitted patients with acute pancreatitis were studied. Bacterial DNA was detected in six patients (19.3%), and the sequencing process allowed identification of Citrobacter freundii and Pseudomonas aeruginosa. In two patients the same bacteria detected at admission was detected 24 hours later (above 99.9% homology of nucleotide sequence). Basic clinical and biochemical characteristics were similar among patients with or without the presence of bacterial DNA. CONCLUSION: Detection of gram negative bacteria derived bacterial DNA in our series supports the contention that bacterial translocation is a systemic process in approximately 20% of patients with acute pancreatitis that does not seem to be related to the severity of the episode or immediate development of infection.

Acute Disease↗
PubMed2006

Crucial role of the melanocortin receptor MC1R in experimental colitis.

BACKGROUND AND AIMS: alpha-Melanocyte stimulating hormone (alpha MSH) is known to exert anti-inflammatory effects, for example in murine DSS (dextran sodium sulphate induced) colitis. The anti-inflammatory functions of alpha MSH are mediated by the melanocortin1-receptor (MC1R) in an autoregulatory loop. The aim of this study was therefore to determine whether a breakdown of the alpha MSH-MC1R pathway leads to worsening of disease. METHODS: Experimental colitis was induced in mice with a frameshift mutation in the MC1R gene (MC1Re/e), C57BL/6 wild type mice, and MC1Re/e-C57BL/6 bone marrow chimeras. The course of inflammation was monitored by weight loss, histological changes in the colon, and myeloperoxidase activity. In addition, MC1R expression was analysed in intestinal epithelial cells. RESULTS: While the colon of untreated MC1Re/e appeared normal, the course of DSS-colitis in MC1Re/e mice was dramatically aggravated, with a significantly higher weight loss and marked histological changes compared to C57BL/6WT. The inflammation eventually led to death in all MC1Re/e, while all C57BL/6WT survived. Similar observations were detected in a transmissible murine colitis model induced by Citrobacter rodentium. Infected MC1Re/e showed delayed clearance of infection. To determine whether missing haematopoietic cell expressed MC1R was responsible, DSS colitis was induced in MC1Re/e-C57BL/6 bone marrow chimeras. MC1Re/e mice receiving MC1R+ bone marrow showed a similar course of inflammation to non-transplanted MC1Re/e. Likewise, transplantation of MC1R bone marrow into C57BL/6WT mice did not lead to any worsening of disease. CONCLUSIONS: This is the first study to show a functional role of MC1R in intestinal inflammation. The data suggest a pivotal role of non-haematopoietic cell expressed MC1R in the host's response to pathogenic stimuli.

Animals↗
PubMed1978

Some interesting isolates from a diagnostic laboratory.

Citrobacter koseri, Plesiomonas shigelloides, Edwardsiella tarda, Yersinia enterocolitica, Alkalescens dispar, Vibrio parahaemolyticus, and Vibrio alginolyticus were seven interesting microorganisms isolated recently in our diagnostic laboratory.

Bacteria↗
PubMed1983

Phenomenon of resistance to Augmentin associated with sensitivity to ampicillin: occurrence and explanation.

Ampicillin was found to be some tenfold more active than amoxycillin against Enterobacter cloacae. This finding explains the observation that some Ent cloacae strains are sensitive to ampicillin in the disc test but resistant to Augmentin. Ampicillin was also found to be more active than amoxycillin against Citrobacter freundii and Serratia marcescens. In view of these findings, the practice of using ampicillin discs to predict sensitivity to amoxycillin should be reconsidered. The use of both ampicillin and amoxycillin discs is appropriate if errors are to be avoided.

Amoxicillin↗
PubMed1993

Use of 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside for the isolation of beta-galactosidase-positive bacteria from municipal water supplies.

A new medium, mX-Gal, has been developed for the membrane filter enumeration of beta-galactosidase-positive bacteria in municipal water supplies. mX-Gal medium contains the chromogenic beta-galactosidase substrate 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal). All Aeromonas, Citrobacter, and Enterobacter strains isolated from raw water on mX-Gal medium were beta-galactosidase positive. In contrast, only 10 to 20% of these strains produced a red colony with a metallic sheen on m-Endo agar LES medium. Of 674 chlorinated water samples analyzed for total coliforms on m-Endo agar LES medium and for beta-galactosidase-positive bacteria on mX-Gal medium, 18 that were negative for coliforms on m-Endo agar LES showed beta-galactosidase-positive bacteria on mX-Gal. Of a total of 50 beta-galactosidase-positive bacteria isolated from these samples, 76% were identified as Aeromonas hydrophila.

Aeromonas↗
PubMed2006

Dual alterations in casein kinase I-epsilon and GSK-3beta modulate beta-catenin stability in hyperproliferating colonic epithelia.

Casein kinase I (CKI)-epsilon and GSK-3beta phosphorylate beta-catenin at Ser(45) (beta-cat(45)) and Thr(41)/Ser(37,33) (beta-cat(33,37,41)) residues, thereby facilitating its ubiquitination and proteasomal degradation. We used a Citrobacter rodentium-induced transmissible murine colonic hyperplasia (TMCH) model to determine Ser/Thr phosphorylation and biological function of beta-catenin during crypt hyperproliferation. TMCH was associated with 3-fold and 3.3-fold increases in CKI-epsilon cellular abundance and 2-fold and 1.8-fold increase in its activity at 6 and 12 days after infection, respectively. beta-Catenin coimmunoprecipitated with both cellular and nuclear CKI-epsilon and cellular axin at these time points. Cellular beta-catenin was constitutively phosphorylated at Ser(45) and underwent subcellular redistribution to cytoskeletal and nuclear fractions at days 6 and 12 of TMCH, respectively. beta-cat(33,37,41), however, exhibited only subtle changes in either phosphorylation status or subcellular distribution even after blocking proteasomal degradation in vivo. Interestingly, GSK-3beta underwent increased phosphorylation at Ser(9), leading to 40% and 70% decreases in its activity at these time points, respectively. Coimmunoprecipitation studies exhibited strong association of GSK-3beta with PKC-zeta at either time point. Cellular beta-cat(45) stabilized and, along with unphosphorylated beta-catenin, underwent nuclear translocation, associated with nuclear accumulated Tcf-4 and cAMP response element binding protein binding protein, and was significantly acetylated, leading to increases in DNA binding. Priming of beta-catenin at Ser(45) exists in vivo. However, beta-cat(45) does not necessarily enter the degradation pathway. Impairment in linking beta-cat(45) to subsequent GSK-3beta-mediated phosphorylation and degradation may account for increased steady-state levels of both unphosphorylated as well as Ser(45)-phosphorylated beta-catenin, which may be causally linked to increases in cell census during TMCH.

Animals↗
PubMedDate not supplied

Silver nitrate: antimicrobial activity related to cytotoxicity in cultured human fibroblasts.

The aims of this study were to ascertain whether silver nitrate (AgNO3) concentrations below those used in clinical practice inhibit bacterial growth, and in parallel study the cytotoxic effects on human fibroblasts. The cytoprotective effects of fetal calf serum (FCS) were also evaluated. The cytotoxic effects of eight different silver nitrate concentrations were determined by assessing mitochondrial activity of viable cells capable of cleaving tetrazolium salts. Antimicrobial activity of AgNO3, range: 7-550 x 10(-5)%, was tested against Staphylococcus aureus, Citrobacter freundii, and Pseudomonas aeruginosa. Silver nitrate concentrations exerting antimicrobial effects were: S. aureus, >70 x 10(-5)%; P. aeruginosa, >/=270 x 10(-5)%, and C. freundii, >/=550 x 10(-5)%. With 2% FCS, the lowest AgNO3 concentration studied (7 x 10(-5)%) showed cytotoxic effects (cell survival 71 +/- 19%) at only 2 h of incubation. Under these conditions AgNO3 cytotoxicity was time- and concentration-dependent in all exposure periods. Cytotoxicity was greatly enhanced causing 76% fibroblast growth inhibition at concentrations of 14 x 10(-5)% and contact time of 2 h. The AgNO3 concentration of 7 x 10(-5)% was also cytotoxic with 5% FCS in the media compared with controls, although cell survival was higher than with 2% FCS. The cytoprotective action of FCS was clearly shown at the concentration of 10% at which AgNO3 cytotoxicity of 7 x 10(-5)% to 28 x 10(-5)% was partially or completely inhibited. Our results show that AgNO3 at concentrations 100-700 times more diluted than that normally used in clinical practice retained effective inhibitory activity against some of the above-mentioned microorganisms. However, even these concentrations are cytotoxic for cultured fibroblasts. Thus, silver nitrate concentrations up to 100 times more diluted can be used, since they possess bacterial growth-inhibiting power, are less cytotoxic and therefore favour wound healing.

Anti-Infective Agents, Local↗
PubMed1985

Inhibitory activity of cefpiramide on beta-lactamase.

The beta-lactamase stability and inhibitory activity of cefpiramide were investigated. Cefpiramide was found to be stable to hydrolysis and inhibited beta-lactamase produced by Citrobacter freundii, Enterobacter cloacae, Morganella morganii, and Escherichia coli. Kinetic studies showed that cefpiramide is a competitive inhibitor of cephaloridine hydrolysis by E. cloacae beta-lactamase.

Cefotaxime↗
PubMed1985

The interaction of beta-lactam compounds with chromosomally mediated enzymes: relations to the molecular structure.

The interaction of 25 beta-lactam compounds with both chromosomally mediated beta-lactamases from Enterobacter cloacae and Citrobacter freundii was studied by enzyme kinetics. All the penams, cephems, and the penem Sch 29482 revealed 'competitive inhibition' of both enzymes. The penem required a preincubation period of approximately 5 min before reaching a state of equilibrium between the active and the inactive enzyme. Except for the recently developed compound HR 810, newer cephalosporins generally exhibited a high affinity for both enzymes. Consistent with earlier findings latamoxef, N-formimidoyl thienamycin, aztreonam and clavulanic acid showed a time dependent inhibition of the enzyme activity. These findings suggest a more complex reaction scheme including a second reversible complex. The last-mentioned compounds share one structural property: the modification or even the loss of the ring fused to the beta-lactam ring. Among these compounds, clavulanic acid exhibited the lowest affinity for the enzymes. It seems likely that the affinity of beta-lactam compounds is mainly influenced by the nature of the substituents.

Anti-Bacterial Agents↗
PubMed1990

Role of the outer membrane for quinolone resistance in enterobacteria.

Quinolone-resistant clones were selected from clinical Escherichia coli, Citrobacter freundii and Serratia marcescens isolates in a frequency ranging from 10(-8) to 10(-6). The outer membrane proteins of quinolone-resistant E. coli clones remained unaltered, as was the case for 10 of 11 C. freundii and 4 of 11 S. marcescens clones ('nal B' type). There was no strong relation between alterations of outer membrane proteins and cross-resistance with chemically unrelated compounds such as tetracycline or chloramphenicol; however, tetracycline resistance was observed in some C. freundii clones with unaltered outer membrane proteins ('mar A'). Most of the quinolone-resistant S. marcescens clones can be considered 'nor B' or 'nor C' mutants due to their cross-resistance with other compounds, their altered outer membrane proteins and changes of lipopolysaccharide. In a few cases, subinhibitory quinolone concentrations caused alterations of outer membrane proteins in S. marcescens during mid log phase without development of resistance.

Anti-Infective Agents↗
PubMed1978

Transmissible murine colonic hyperplasia.

After exposure to a variant of Citrobacter freundii, suckling and adult mice developed transmissible murine colonic hyperplasia of the same degree of severity. Mucosal hyperplasia was most severe 2 to 3 weeks after inoculation and then regressed. Suckling mice had a high mortality because of secondary inflammatory and erosive changes. Severe hyperplasia was characterized by mitotic activity along the entire crypt column and surface mucosa.

Animals↗
PubMed2006

Carriage by the German cockroach (Blattella germanica) of multiple-antibiotic-resistant bacteria that are potentially pathogenic to humans, in hospitals and households in Tripoli, Libya.

Using standard bacteriological procedures, 403 cockroaches (Blattella germanica) collected in Tripoli, from hospitals or the households surrounding the hospitals, were examined for bacteria that are potentially pathogenic to humans. Almost all of the cockroaches (96.1% of the 253 from hospitals and 98.7% of the 150 from households) were found to be carrying potentially pathogenic bacteria, with similar mean burdens of 3.2 x 10(5) colony-forming units (cfu) (range=0-1.4 x 10(7)) for each hospital cockroach and 1.9 x 10(5) cfu (range=0-3.1 x 10(6)) for each household cockroach (P>0.05). Overall, 27 and 25 species of potential pathogen were isolated from the hospital and household cockroaches, respectively, with Klebsiella, Enterobacter, Serratia and Streptococcus predominant. Carriage of species of Serratia was significantly more common among the hospital cockroaches than among the household cockroaches, whereas carriage of Klebsiella, Enterobacter, Citrobacter and Aeromonas was significantly more common among the household cockroaches than among the hospital. Multiple resistance, to at least six different antibiotics, was more commonly observed among the enteric bacteria isolated from the hospital cockroaches than among those recovered from the household cockroaches. Overall, >30% of the isolates of Enterobacteria recovered were each resistant to at least four antimicrobial agents, and 95% of the Pseudomonas isolates were each resistant to at least eight such agents. Cockroaches may play an important role in the spread of multiple-antibiotic-resistant, bacterial pathogens within the hospitals and surrounding communities of Tripoli and other, similar cities. The local health and environmental authorities need to be encouraged to treat B. germanica infestations seriously and to control them quickly and effectively.

Aeromonas↗
PubMed2006

Animal models of inflammatory bowel disease: lessons from enteric infections.

Mouse models of intestinal inflammation have played a key role in understanding the mechanisms that govern the inflammatory response in the intestine, and in designing new therapeutic strategies in the treatment of patients with inflammatory bowel disease (IBD). Most of these models use chemical challenges, whereas relatively few robust models of intestinal inflammation caused by microbial infection are known. Two common models of infectious murine colitis and typhlitis are infection with the murine epithelial-adherent pathogen, Citrobacter rodentium, and infection of streptomycin-pretreated mice with Salmonella typhimurium. Studies in these models have helped to define the interactions between bacterial pathogens and host immune defenses, thus broadening the understanding of host-microbial interactions in the intestinal tract. Furthermore, such models help to determine the physiologic consequences of neutralizing specific mediators and signaling pathways implicated in inflammation on antimicrobial host defense.

Animals↗
PubMed1998

Analysis of enterohemorrhagic Escherichia coli serotype O157:H7 by flow cytometry using monoclonal antibodies.

To develop a rapid and specific method to detect and/or identify enterohemorrhagic Escherichia coli O157:H7, two mouse monoclonal antibodies (MAbs) were prepared. Specificities of these two MAbs (1D9 and 3E8) were determined by flow cytometry method (FCM). MAbs 3E8 and 1D9 were found to react with E. coli O157:H7, Citrobacter freundii and Salmonella group N (O:30), but not with Escherichia hermannii. With a mixture containing strains of E. coli O157:H7 and E. coli O6:H1, two different peaks appeared in FCM with MAbs, whereas a single peak appeared with polyclonal rabbit antiserum. From these findings, FCM with MAb is suggested to be a rapid, specific, and useful method to detect and identify strain(s) of E. coli O157:H7 in food ingredients.

Animals↗