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Citrobacter meningitis in adults.

Citrobacter meningitis is an uncommon infection of neonates and young children. It is rarely seen in adults. We describe a 46-year-old man with a mixed bacterial meningitis caused by C. diversus and Klebsiella oxytoca and a 64-year-old woman with C. freundii meningitis. Review of the English-language literature revealed only 2 adult patients with C. diversus meningitis and another 2, with C. freundii meningitis. The ages of these 6 aforementioned patients ranged from 31 to 84 years. Multiple facial fractures, neurosurgical procedures, alcoholism and diabetes mellitus were predisposing conditions. Among the 5 patients whose outcome was known, antibiotic therapy was successful in 4 but failed in 1. This study emphasizes that almost any of the gram-negative bacilli can cause serious infection of the central nervous system in adults in the proper setting.

Adult↗

[The use of random amplification of polymorphous bacterial DNA for the typing of bacteria in the genus Citrobacter].

To obtain the profiles of randomly amplified DNA, isolated from bacteria of the genus Citrobacter, the method of polymerase chain reaction was used. Nine oligonucleotides were evaluated for the possibility of their use as primers for the amplification of random polymorphous sequences of DNA; of these, 2 oligonucleotides which generated profiles, sufficiently reproducible and typical for different C. freundii and C. diversus strains, were selected. The possibility of using the above oligonucleotides in pair for amplification of species-specific fragments of polymorphous bacterial DNA for typing was shown.

Base Sequence↗

A large nontypical outbreak of Norwalk virus. Gastroenteritis associated with exposing celery to nonpotable water and with Citrobacter freundii.

The US Air Force Academy experienced a point-source outbreak of gastroenteritis originally believed to be caused by Salmonella. The overall attack rate was 48% among approximately 3000 cadets and staff. Food-specific attack rates implicated chicken salad. The odds ratio for chicken salad consumption in ill cadets was 10.7 (95% confidence interval: 8.2; 13.8). The celery component had been exposed to nonpotable water. Citrobacter freundii were statistically associated with consumption of the suspected vehicle and subsequent illness. Most aspects were consistent with the epidemiology of Norwalk gastroenteritis. However, the clinical presentation was not typical of reported outbreaks. One hundred five cadets required intravenous rehydration. Serum samples implicated Norwalk virus as the most probable cause of this outbreak. The Centers for Disease Control (Atlanta, Ga) recently began national surveillance for viral gastroenteritis. All outbreaks of gastroenteritis associated with nonpotable water should be investigated for evidence of viral cause.

Citrobacter freundii↗

Phosphate release and heavy metal accumulation by biofilm-immobilized and chemically-coupled cells of a Citrobacter sp. pre-grown in continuous culture.

A heavy metal-accumulating Citrobacter sp. was grown in carbon-limiting continuous culture in an air-lift fermentor containing raschig rings as support for biofilm development. Planktonic cells from the culture outflow were immobilized in parallel on raschig rings by chemical coupling (silanization), for quantitative comparison of phosphatase activity and uranyl uptake by both types of immobilized cell. The flow rate giving 50% conversion of substrate to product (phosphate) in flow-through reactors was higher, by 35-40%, for the biofilm-immobilized cells, possibly exploiting a pH-buffering effect of inorganic phosphate species within the extracellular polymeric material. Upon incorporation of uranyl ions (0.2 mM UO22+), both types of cell removed more than 90% of the input UO22+ at slow flow rates, but the chemically-coupled cells performed better at higher flow rates. The deposited material (HUO2PO4) subsequently removed Ni2+ from a second flow via intercalative ion exchange of Ni2+ into the crystalline HUO2PO4.4H2O lattice. This occurred irrespective of the method of coupling of the biomass to the support and suggested that uranyl phosphate accumulated by both types of cell has potential as a bio-inorganic ion exchanger-a potential use for the uranium recoved from primary waste treatment processes.

Animals↗

Neonatal Citrobacter meningitis: pathogenesis of cerebral abscess formation.

In 2 premature infants with brief survival, the major pathological feature of Citrobacter meningitis consisted of vasculitis and infarction with necrosis and liquefaction of large portions of the white matter of the hemispheres. Areas of decreased density demonstrated on computed tomographic head scan represented infarction with necrosis, liquefaction, and cavitation and not a classic abscess. This finding has clinical significance, because surgical drainage of liquefied infarcts is rarely indicated and could further damage the relatively preserved cortex.

Brain Abscess↗

Isolation of Citrobacter sp. mutants defective in decolorizing malachite green.

To identify genes involved in the decolorization of malachite green, random mutants generated by transposon insertion in the malachite green-decolorizing bacterium, Citrobacter sp. were isolated. The resulting mutant bank yielded 24 mutants with complete defects in their abilities to decolorize malachite green. Southern hybridization with a Tn5 fragment as a probe showed a single hybridized band in 7 mutants, which appeared to have insertions at different sites of the chromosome. The Tn5-inserted genes were isolated and the DNA sequence flanking Tn5 was determined. Based on a sequence database, the putative protein products encoded by the mg genes were identified as follows. mg3, an ABC transporter homolog; mg6, a LysR-type regulatory protein; m11, an oxidoreductase; mg17, a MalG protein in the maltose transport system; and mg21, a sugar kinase. The deduced sequences from two mg genes (mg7 and mg18) showed no significant similarity to any protein with a known function, suggesting that these two mg genes encode unidentified proteins that are responsible for the decolorization of malachite green.

Bacterial Proteins↗

Cloning and sequencing of the gene for the lactose carrier of Citrobacter freundii.

The gene coding for the lactose carrier of Citrobacter freundii was cloned into the plasmid pBR322. The gene was sequenced and the amino acid sequence was found to be 70% identical to the lactose carrier of E. coli. All of the charged residues in the membrane spanning region were conserved. The sugar specificity is somewhat different from that of E. coli. The C. freundii carrier has less activity for lactose and more activity for o-nitrophenyl-galactose (ONPG) than the carrier of E. coli.

Amino Acid Sequence↗

Crystal structure of Citrobacter freundii restriction endonuclease Cfr10I at 2.15 A resolution.

The X-ray crystal structure of Citrobacter freundii restriction endonuclease Cfr10I has been determined at a resolution of 2.15 A by multiple isomorphous replacement methods and refined to an R-factor of 19.64%. The structure of Cfr10I represents the first structure of a restriction endonuclease recognizing a degenerated nucleotide sequence. Structural comparison of Cfr10I with previously solved structures of other restriction enzymes suggests that recognition of specific sequence occurs through contacts in the major and the minor grooves of DNA. The arrangement of the putative active site residues shows some striking differences from previously described restriction endonucleases and supports a two-metal-ion mechanism of catalysis.

Binding Sites↗

Anaerobic malonate decarboxylation by Citrobacter diversus. Growth and metabolic studies, and evidence of ATP formation.

Citrobacter diversus ATCC 27156 was able to grow by decarboxylation of malonate to acetate under strictly anaerobic conditions, in the presence of yeast extract. The growth yield, corrected for growth on yeast extract, was 2.03 g cell dry mass per mol malonate. The addition of malonate to ATP-depleted cell suspensions (less than 0.2 nmol ATP/mg cell protein) resulted in a rapid increase in cellular ATP levels to between 4.5 and 6.0 nmol/mg cell protein. Intact cells decarboxylated malonate at rates of up to 1.5 mumol/min.mg protein. Enzyme assays on malonate-grown cells indicated activation of malonate by an ATP-dependent ligase reaction and by CoA transfer from acetyl-CoA, followed by decarboxylation of malonyl-CoA to acetyl-CoA with subsequent recovery of the invested ATP by substrate level phosphorylation through the activity of acetate kinase. Net ATP synthesis is postulated to be mediated by gradient formation coupled to the decarboxylation of malonyl-CoA. The protonophore CCCP and H(+)-ATPase inhibitor DCCD significantly reduced cellular ATP levels, suggesting a role for proton gradients in the energy metabolism of this strain when growing an malonate. Inhibitors of sodium metabolism or ommission of sodium had no effect on ATP levels or malonate decarboxylation.

Adenosine Triphosphate↗

Cloning and expression of the gene(s) for cephalosporinase production of Citrobacter freundii.

The chromosomal gene(s) for cephalosporinase production of Citrobacter freundii GN346 has been cloned into vector plasmid pMK1, initially as a 7.3 kb EcoRI fragment. From the substrate profile and the response to anti-GN346 CSase serum of the enzyme produced, it was confirmed that the hybrid plasmid (pTY71) carries the relevant chromosomal cephalosporinase gene from C. freundii GN346. A restriction endonuclease cleavage map of cloned EcoRI fragments was constructed, and the structural gene of the cephalosporinase could be limited in the 1.5 kb BamHI fragment. The cloned gene(s) was expressed at an extremely low level in Escherichia coli. Furthermore, its expression was constitutive in E. coli, although inducible in its own cytoplasm.

Cephalosporinase↗

Growth conditions and heat resistance of Citrobacter freundii.

The influence of the growth medium and the growth temperature on the heat resistance of Citrobacter freundii has been established. Logarithmic growth phase cells grown on rich media have a higher heat resistance than cells of the same phase grown on minimal media. This finding was independent of type of carbon source in the growth medium, but the kind of carbon source has a definite influence on the heat resistance. Logarithmic phase cells grown at 37 degrees C are much more heat stable than cells grown at 20 or 41 degrees C. Stationary growth phase cells are much more heat resistance than logarithmic phase cells, whereas Mg2+ - or glucose-starved cells are even slightly more heat stable than stationary phase cells.

Bacterial Proteins↗

Formate dependent nitrate and nitrite reduction to ammonia by Citrobacter freundii and competition with denitrifying bacteria.

Citrobacter freundii, Paracoccus denitrificans and Pseudomonas stutzeri were grown either singly or in mixed culture in anaerobic nitrate or nitrite limited chemostats with formate and/or succinate as electron donors and carbon sources. C. freundii reduced nitrate or nitrite stoichiometrically to ammonia. Maximum molar growth yields for nitrate (nitrite) were 15.3 (9.9) g/mol for C. freundii on formate with succinate as carbon source, 15.3 (9.5) g/mol for Ps. stutzeri on succinate and 32.3 (20.4) g/mol for Pa. denitrificans on succinate. The almost identical growth yields indicate that the ATP output of the anaerobic processes in the nitrate (nitrite) ammonifying organism and Ps. stutzeri are nearly the same. In mixed cultures with either Ps. stutzeri or Pa. denitrificans, C. freundii was the best competitor for nitrate. These results show that in anaerobic environments C. freundii may compete successfully with denitrifying organisms.

Ammonia↗

Selection of resistant mutants of Citrobacter freundii by second and third-generation cephalosporins and imipenem.

Using a single-step selection procedure, resistant mutants could be obtained from three clinical isolates of Citrobacter freundii with two second-generation and four third-generation cephalosporins but not with imipenem. All mutants showed a drastically increased beta-lactamase activity and were cross-resistant to all the cephalosporins examined. Combinations of cloxacillin with the cephalosporins were markedly synergistic, suggesting the principal role of the cephalosporinase in the resistance of these mutants.

Cephalosporins↗

Evidence for nonspecific induction of beta-lactamase in overproducing variants of Enterobacter cloacae and Citrobacter freundii.

Induction of chromosomally mediated beta-lactamases was studied in clinical isolates of Enterobacter cloacae and Citrobacter freundii. Whereas isolates resistant to ampicillin and cefoxitin exhibited an inducible enzyme, those sensitive to both agents did not. Cefoxitin and above all imipenem proved the most efficacious enzyme inducers. Variants among these inducible isolates which produced large amounts of beta-lactamase could be selected in the presence of cefamandole. In each of these selected variants constitutive enzyme production was markedly enhanced by induction, even with compounds lacking induction potency for the corresponding wild strains. Moreover, these variants could also be induced non-specifically, i.e. by non-beta-lactam compounds. Production of large amounts of enzyme was associated with resistance to all available beta-lactam compounds.

Anti-Bacterial Agents↗

Genetics of colicin E susceptibility in citrobacter freundii.

The insensitivity of Citrobacter freundii to the E colicins is based on tolerance to colicin E1 and resistance to colicins E2 and E3. Spontaneous colicin A resistant mutants of C. freundii also lost their colicin E1 receptor function. Sensitivity to colicin E1 can be induced by F'gal+tol+ plasmids, the tol A+ gene product of which is responsible for this effect. Receptor function for colicins E2 and E3 is induced by the E. coli F'14 bfe+ plasmid, which is also able to enhance notably the receptor capacity for colicin E1. The bfe+ gene product of E. coli, which is responsible for these phenomena, also restores the receptor function for colicin A and E1 in colicin A resistant mutants of C. freundii. All results show that there is a remarkable difference between the E. coli bfe+ gene product and the bfe+ gene product of C. freundii and also between the tol A+ gene products of these strains. The sensitivity to phage BF23 parallels the sensitivity to colicins E2 and E3 and is also induced by the F'14 bfe+ plasmid.

Citrobacter↗

A genetic study of tolerance and resistance to colicin A in Citrobacter freundii.

Colicin A-insensitive mutants of Citrobacter freundii were isolated and grouped into six phenotypic classes characterized by sensitivity, insensitivity or partial insensitivity to the bacteriocins S6, DF 13 and colicin A, and sensitivity or insensitivity to deoxycholate (DOC) and ampicillin. Mapping by the gradient-of-transmission method revealed the chromosomal regions in which the responsible genes are situated. Res-3 mapped near pur between pur and thr; Tol-5 mapped between aro and ilv and Tol-4 between gal and pur; Tol-1, Tol-2 and Tol-3 are situated close to gal. All the mutations that mapped near gal rendered the bacteria more sensitive to DOC and ampicillin. Complementation analysis with E. coli plasmids showed that the three phenotypic groups that map near gal were complemented by E. coli plasmids and fall into three complementation groups. Two of these are quivalent with the tol A and tol B genes in E. coli.

Chromosome Mapping↗

Basic studies of hydrogen evolution by Escherichia coli containing a cloned Citrobacter freundii hydrogenase gene.

Citrobacter freundii genes that complemented Escherichia coli hyd-(hydrogenase activity) mutation were cloned in plasmids pCBH4 (6.2 kb) and pCBH6(5.7 kb). Hydrogen evolution by the transformant E. coli HK-8(pCBH4 or pCBH6) was investigated. The optimum culture temperature of recombinant E. coli cells for hydrogen evolution from glucose was in the neighborhood of 18 degrees C. The recombinant E. coli cells cultured at this condition showed a several-fold increase of hydrogen evolution, as compared with that of the wild-type cells. The plasmid-retention stability of this recombinant E. coli was extremely high, especially plasmid pCBH4, which was completely retained during 2 wk without any restriction. Hydrogen production by immobilized recombinant E. coli was then investigated using cells cultured at 18 degrees C. The hydrogen evolution rate from glucose and Lennox-broth were about twofold higher than that of E. coli C600, and this high hydrogen evolution rate was maintained for more than 1 mo.

Citrobacter↗

Citrobacter sepsis in infants.

A study of blood cultures from 320 cases of neonatal sepsis showed 136 (42.5%) to be positive for bacterial growth; of these 82 (60.29%) isolates being gram negative bacilli. Citrobacter was the commonest gram negative bacilli isolated. Other commonly isolated gram negative organisms were Pseudomonas, Klebsiella, Salmonella typhimurium, Acinetobacter and Escherichia coli. Antibiotics susceptibility pattern revealed the isolates to be resistant to commonly used antibiotics.

Bacteremia↗