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[Clinical and experimental studies on the pathogenicity of clinical isolated Citrobacter freundii].

Clinical and fundamental studies were performed using Citrobacter freundii isolated from urine specimens of in- and out-patients between January 1980 and July 1981. The C. freundii (greater than 10(4)/ml) isolated from urine was closely related to chronic complicated urinary tract infection. MICs of pyridonecarbonic acids (ENX, OFLX and NFLX) and CMX against C. freundii were low. Pyridonecarbonic acids (ENX, OFLX and NFLX) and CMX showed good therapeutic effects against the experimental ascending pyelonephritis of mice according to their MICs. Against the experimental ascending pyelonephritis of cyclophosphamide-treated mice, pyridonecarbonic acids (ENX, OFLX and NFLX) showed good therapeutic effects. Experimental intraperitoneal infection of mice was made easily with a low inoculation dose by adding mucin. Pyridonecarbonic acids (ENX, OFLX and NFLX) and CMX showed good therapeutic effects against this experimental model. Experimental respiratory tract infection could not be made in the mice with the organism isolated from urine.

Animals↗

[Enzymatic synthesis of 3,4-dihydroxyphenyl-L-alanine by free and immobilized Citrobacter freundii cells].

The Citrobacter freundii 62 cells immobilized in PAAG and possessing the tyrosine-phenol-lyase (TPL) activity catalyse the synthesis of 3,4-dihydroxyphenyl-L-alanine (DOPA) from pyrocatechol and ammonium pyruvate. The synthesis of DOPA was studied using both free and immobilized bacterial cells. When the concentration of pyrocatechol is over 0.1 M the TPL activity of the cells is inhibited. The concentration of pyrocatechol can be increased up to 0.3 M by using an equimolar mixture of pyrocatechol and boric acid. The addition of ascorbic acid as an antioxidant results in a lower TPL activity of both free and immobilized bacterial cells.

Acrylic Resins↗

[Synthesis of tyrosine and 3,4-dihydroxyphenylalanine by Citrobacter freundii bacteria from lactic acid].

A possibility of using lactic acid as a precursor for the synthesis of L-tyrosine and L-3,4-dihydroxyphenylalanine (DOPA) by Citrobacter freundii 62 and 63 was established. The synthesis of tyrosine from lactic acid occurs at a phenol concentration of less than 0.6%. The conditions were found which enable C. freundii 62 and 63 to synthesize from lactic acid tyrosine and DOPA with the yield of 35-38 g/l and 32-33 g/l, respectively.

Aerobiosis↗

Toxic exoproducts of citrobacter freundii.

A strain of Citrobacter freundii isolated from the feces of a patient with diarrhoea was examined for growth kinetics and toxic exoproduct formation using the complete (BHI) and synthetic culture media. It was found that the test organism in synthetic medium grew distinctly slower than in BHI. Fractionations on Sephadex G-100 column yielded 3 fractions from the complete medium culture filtrate and 2 fractions from the culture filtrate obtained from synthetic medium. The first culture filtrate fractions (F1) were represented by components of the molecular weight over 100,000, the respective second fractions (F2) from complete and synthetic medium were of the molecular weights of about 40,000 and 10,000. In the early skin test on rabbits the toxicity of culture filtrates and their fractions manifested itself by an increased permeability of blood vessels, in the late skin test by a hemorrhagic reaction associated with dilatation of blood vessels and induration of the skin tissue. In a test on mouse foot pad all separated filtrate fractions gave a positive edematous reaction. In cultured Vero cells samples of synthetic medium fractions gave a distinct cytotoxic reaction. Immunochemically, the presence of LPS in culture filtrates as well as some variations in the antigenicity of components from the complete and synthetic medium fractions were found. Apart from LPS some additional high-molecular-weight components were also present in the toxic complex of both first filtrate fractions (F1). Much more attention should be given to analysis of these first fraction complexes as well as to toxinogenicity of second fractions (F2) using some additional tests.

Bacterial Toxins↗

[Diagnostic value of the acid production from galacturonate to differentiate strains of "Salmonella" sub-genus I and monophasic sub-genus III from the other "Salmonella" strains belonging to sub-genera II, diphasic III, IV, "Citrobacter" and "Hafnia alvei" (author's transl)].

Two methods were compared: (1) the ability to use sodium glacturonate as source of carbon and energy in defined medium; (2) the ability to produce acid by fermentation of this substrate in peptone water with phenol red. Results of growth on, acid formation from, galacturonate are identical. Salmonella strains of sub-genus I and monophasic serotypes of sub-genus III do not use galacturonate. Salmonella of sub-genera II and IV as well as diphasic serotypes of sub-genus III use galacturonate. Glacturonate-negative Salmonella are isolated mainly from man and warm-blooded animals whereas galacturonate-positive Salmonella are most often isolated from cold blooded animals and from the enrivonment. Citrobacter and Hafnia alvei which may be confused with Salmonella spp. are galacturonate-positive.

Citrobacter↗

[Synthesis of tyrosine and 3,4-dihydroxyphenylalanine by bacteria Citrobacter freundii].

Among facultative-anaerobic bacteria utilizing formic acid, a large number of strains having tyrosine phenol lyase were found. The enzyme can catalyze synthesis of tyrosine and 3,4-dihydroxy phenyl alanine (DOPA) from pyruvate, ammonium and, accordingly, phenol and pyrocatechol. These strains were identified as Citrobacter freundii. Cell suspensions of the most active strains synthesized up to 75 g/l tyrosine for 12 hr, up to 86 g/l tyrosine for 24 hr, and up to 29 g/l DOPA for 42 hr. A medium containing yeast autolysate grown on hydrocarbons can be recommended to produce cells having a high tyrosine phenol lyase activity.

Ammonia↗

The effect of 2-oxoglutarate and biotin in the release of amino acids by Citrobacter intermedius C3.

Excretion of amino acids by Citrobacter intermedius C3 was assayed in a mineral medium with glucose as carbon source. Glutamic acid is the main amino acid excreted in liquid medium and it is also detected at the colonial level in solid medium. Mutants with different behaviour with respect to the excretion of amino acids are studied. The presence of 2-oxoglutarate in the medium induced excretion in all strains. On the other hand when biotin was added to the culture media amino acid excretion was partially reduced.

Amino Acids↗

[Monosaccharide composition of the lipopolysaccharides of bacteria of the genus Citrobacter].

The authors studied antigens obtained by Grasset's method from 13 strains of Citrobacter of the International collection. The strains possessed O- and H-antigens whose behaviur in the electric field differed. All the strains under study were divided into two groups (by the number of serologically-active components of their O-antigens); representatives of the second group had no cathode O-antigen component. Chemical composition of specific lipopolysaccharides (LPS) obtained by Westphal's method was determined. Fourteen different sugars were revealed. The strains under study were referred to the known chemotypes. Strain 16/52 (8a, 8c) was for the first time studied in respect to the monosaccharide composition of specific LPS, and was referred to chemotype designated as CC-L.

Agglutination Tests↗

Polysaccharides and their common proteinic carrier in the Vi antigens of Citrobacter ballerup and Salmonella typhi Ty2.

Immunochemical analysis of Citrobacter ballerup and Salmonella typhi Ty2 showed that the strains share native and heat-resistant proteins that are, apparently, the carriers of a common polysaccharidic determinant present in their respective somatic antigens. After the classic acetic acid hydrolysis, the somatic antigen of C. ballerup reacted, in agar gel, against the homologous antiserum by two precipitation lines, one of which also precipitated against the anti S, typhi Ty2 serum; the hydrolysis of the S. typhi Ty2 somatic antigen demonstrated that, in addition to the 'O' polysaccharide, reacting against all the S. typhi antisera, it contains a polysaccharide that precipitated against the anti-C. ballerup serum. The elusiveness in the agglutinability of only freshly isolated bacterial authorizes some doubt concerning the responsibility of the antipolysaccharide antibodies in the agglutinating Vi sera; in order to induce anitpolysaccharides hyperimmunizations are needed while antiproteins are easily induced by short immunizations.

Antigens, Bacterial↗

[TEM-type beta-lactamase coded by the plasmid from Citrobacter sp].

beta-Lactamase was isolated from the cells of E. coli, strain 1039, a transconjugant carrying the plasmid first detected in Citrobacter sp. beta-Lactamase was purified to obtain a homogeneous preparation. The activity of the enzyme was estimated by a modification of the potentiometric method providing determination of up to 0.1 mumol of penicilloinic acid. The activity of the pure enzyme was 206 mumol per 1 mg of protein per minute. The molecular weight determined by gel filtration was 21500. The substrate profile of beta-lactamase corresponded to the TEM type. Preliminary incubation with methicillin resulted in a significant decrease in the enzyme activity. Carbenicillin, dicloxacillin, oxacillin and cephalothin induced no decrease in the activity of the enzyme when subjected to preliminary incubation with it. The use of the functional group reagents allowed detecting residues of serine and lysine in the enzyme active centre or close to it. Km and Kcat for some antibiotics were evaluated. The ratio of alpha- and beta-structures in the molecule of beta-lactamase was determined with the method of circular dichroism (CD). The fraction of the alpha-spiral areas amounted to 30 +/- 5 per cent and the fraction of beta-structures amounted to 20 +/- 5 per cent. On attachment of methicillin to the molecule of the enzyme the ratio of alpha- and beta-structures did not change which may be considered as a preliminary indication of the centre "immobility" in the process of catalysis. Slow hydroysis of methicillin by beta-lactamase was shown with the method of CD.

Chemical Phenomena↗

[Comparison of antibacterial potencies of oral and parenteral antibiotic preparations against Escherichia coli, Klebsiella, Citrobacter, and Proteus isolated from urinary tract infections (3: 1981). 2. Changes in bacterial sensitivities].

Since 1979 the antibacterial activity of antibiotics against E. coli, Klebsiella, Citrobacter and Proteus isolated from patients with urinary tract infections has been investigated. The serious transition of susceptibilities of E. coli and Klebsiella could not be recognized in these antibiotics (MPC, ABPC, NA, PPA, CEX, CEZ, CTM, CMZ and CFX). However, a few resistant organisms against the third generation's antibiotics (CTX, CMX, CZX, LMOX and CPZ) have already been appeared, we have to observe these results, continuously.

Anti-Bacterial Agents↗

[Enzyme activity of the formate hydrogenlyase complex in Citrobacter freundii].

Citrobacter freundii 62 can grow in the absence of oxygen in media containing glucose, peptone, fumarate or malate. When the medium contained fumarate or malate, the culture could grow under anaerobic conditions only in the presence of molecular hydrogen, formate or nitrate. The highest activity of formatehydrogenlyase and hydrogenase was found when C. freundii grew in a medium with glucose and formate. The activity was lower in media with other organic substrates, particularly, in the absence of formate or H2. The activity of hydrogenase was very low in cells grown under aerobic conditions or in the presence of nitrates while the activity of formatehydrogenlyase was not found at all for all practical purposes. The activity of formate dehydrogenase assessed in the presence of methylene blue was rather high irrespective of the conditions under which the culture was grown. However, when the activity of formate dehydrogenase was determined in the presence of benzyl viologen, it was high only in cells grown in the medium with glucose and formate.

Aerobiosis↗

[The GC content of a group of H2S- enterobacterial related to the genus Citrobacter].

The deoxyribonculeic acid (DNA) of 106 strains of Enterobacteria was analysed for the guanine + cytosine (GC) content. These strains, whose origin and principal characters are described in the text, belong to the genera Citrobacter (C. freundii H2S-) and Levinea (L. malonatica and L. amalonatica). Four other groups or classes named C.D.E. and F. could not be classified on the base of the usual phenotypic criteria. DNA from the strains of Levinea has a GC% of 50.3 to 53.3, while DNA from the strains of C. freundii H2S- has a GC% of 48.6 to 51.7. The representative values from the new classes are C, 50.9%; D, 54%; E, 52.7%; F, 49.5%. For the latter a genomic heterogeneity was shown, expressing itself as two subpopulations whose average GC% are 51.7 and 48.6 respectively. Statistical analysis of the averages give a significant individuality to these new classes.

Citrobacter↗

Complications of Citrobacter neonatal meningitis: assessment by real-time cranial sonography correlated with CT.

Real-time cranial sonography via the anterior fontanelle was used serially over a 3- and 6-week period, respectively, to evaluate two infants who developed multicystic encephalomalacia secondary to Citrobacter neonatal meningitis. Sonographic findings included heterogeneous parenchymal echogenicity, gyral prominence, periventricular hypoechoic areas from which cystic spaces evolved, and development of hydrocephalus. Serial cranial computed tomography over the same time period confirmed the sonographic observations in each case.

Citrobacter↗

Citrobacter koseri isolated in Israel, 1972-83.

Cultures of Citrobacter koseri (syn. C. diversus), an opportunistic pathogen that was isolated in Israel during 1972-83, were studied by sero- and biotyping and for susceptibility to antibacterial agents. Of 1,172 cultures, 96% were isolated from adults, more than half from urinary tract infections. Other frequent sources were superficial wounds and discharges from the respiratory and genital tracts. Seventeen strains originated from blood cultures of patients with sepsis and four from the cerebrospinal fluid of newborns with meningitis. The isolates belonged to 17 O groups, 60 serotypes and 5 biotypes. The most frequent serotypes were 15:a:[2], first identified in Israel, and 1:a:[2]. The most frequent biotypes were c and a. No correlation was found between sero- or biotypes and source of isolation or type of disease. Of 720 cultures tested for sensitivity, all were resistant to ampicillin and carbenicillin, but only 9.3% were resistant to one or more of nine other antibacterial agents tested.

Adult↗

[Immobilization of Citrobacter freundii cells with tyrosine phenol-lyase activity by entrapment in natural gels].

A comparative study on immobilization of Citrobacter freundii cells by entrapment in carrageenan, agar, agar-agar, and gelatin gels (5, 10, and 15%) was carried out. Gelatin gels were treated with glutaraldehyde to make them more rigid. As a result, the tyrosine phenol-lyase activity of these samples was less than that of free cells (about 40%). The yield of TP-lyase activity was 40--60% when cells were immobilized in 5% and 7% agar and agar-agar gels. The cells entrapped in carrageenan gels, the concentration of which was varied from 2 to 10%, possessed the highest tyrosine phenol-lyase activity (up to 90%). The efficiency of cell entrapment was high for all the carrier and equal to 70--90%. The plastic strength and swelling of the above gels, as well as the phenol adsorption on the carriers and the release of the bacterial cells from them were studied under the conditions of tyrosine phenol-lyase reaction.

Adsorption↗

[Isolation and properties of tyrosine phenol-lyase from Citrobacter intermedius].

A method for preparation of homogeneous tyrosine phenol lyase (EC 4.199.2) from Citrobacter intermedius has been developed. The cells were cultivated in the media with a view to obtain a cell culture with a high activity of tyrosine phenol lyase. The isoelectric point for the enzyme lies at pH 4.9. Tyrosine phenol lyase is strictly stereospecific: it catalyzes the formation of pyruvate only from L-tyrosine, but not from D-tyrosine. Kinetic studies showed that K+ and NH4+ cations are non-competitive activators of the enzyme (Ka = 3.57 X 10(-3) and 1.34 X 10(-4) M, respectively).

Chromatography, DEAE-Cellulose↗