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PubMed1980

The effect of acetohydroxamic acid on the induction of bacterial ureases.

The ureases of Proteus mirabilis, Proteus vulgaris, and Proteus rettgeri are inducible by urea. Induction is increased when both urea and acetohydroxamic acid, an inhibitor of urease, are present during bacterial growth. Acetohydroxamic acid alone does not cause induction, but, by preventing the hydrolysis of urea, it minimizes pH increases and allows induction to occur much more effectively. The ureases of Proteus morganii and other bacterial genera are not inducible by this method. The relevance of our findings to the formation and management of infection stones is discussed.

Enzyme Induction↗
PubMed1981

Ureolytic and urease-activating properties of commercial laboratory animal bedding.

Five types of commercial laboratory animal bedding were assessed for endogenous ureolytic activity using a sensitive method which measures the rate of evolution of 14C-carbon dioxide from 14C-urea. On a weight basis, the highest levels of urease activity were found in heat-treated hardwood chips and a regular grade of crushed corncobs. A deodorant-treated type of crushed corncobs had a moderately high level of activity, while pelleted corncobs and pelleted alfalfa were almost devoid of urease activity. A heat-stable activator of bacterial urease was found in hardwood chips and crushed corncobs.

Animals↗
PubMedDate not supplied

[Urease synthesis regulation in Staphylococcus saprophyticus by urea and ammonia].

The effect of growth conditions on urease synthesis was studied with Staphylococcus saprophyticus L-1 isolated from natural sources. Urease biosynthesis was recorded in the absence of urea in the complete medium and in the conditions of nitrogen deficiency; the highest level of the enzyme biosynthesis was found when the culture was grown in the absence of amine nitrogen in the medium. Ammonium ions were a reversible inhibitor of urease and, at a high concentration (30 g of (NH4)2SO4 per litre of the medium), partly repressed its biosynthesis. The rate of growth was low when the cells were cultivated in flasks in a medium containing urea (20 g per litre of the medium). The growth was not inhibited when the cells were cultivated in 20-litre fermenters at the same concentration of urea, but with automatic pH regulation. The alkaline medium rather than urea contained in it appeared to be the principal factor inhibiting growth of the culture.

Ammonia↗
PubMed1995

Confirmation of successful therapy of Helicobacter pylori infection: number and site of biopsies or a rapid urease test.

BACKGROUND: Although a number of tests have been described to detect the presence of Helicobacter pylori in biopsy specimens, studies of positive and negative value have largely been performed on untreated patients; testing the reliability of posttherapy has not been done. METHODS: We examined the value of the number and site of biopsies performed and the method used for specimen evaluation posttherapy. For postantimicrobial therapy of 141 patients with previously confirmed H. pylori infection, three biopsies were taken, two from the antrum and one from the corpus. Individual slides were coded, randomized, and interpreted blindly by two pathologists. Furthermore, in 143 patients, a biopsy specimen was taken from the antrum and was immediately inserted into the gel of the rapid urease test, and the results were compared with those obtained from histopathology obtained at the same time. RESULTS: In 71 patients, H. pylori therapy was unsuccessful; in 61 (86%), all three sites were positive. The highest yield with a single large cup biopsy specimen was 94%; the lowest was 91%. Two antral biopsies were negative in 4% [95% confidence interval (CI) = 1-12%]. The combination of a biopsy from the angulus incisura and one from the greater curvature of the corpus correctly identified all treatment failures (95% CI = 95-100%). The rapid urease test was false-negative in 5% (95% CI = 1-13%); there were no false-positives. CONCLUSION: Use of either the rapid urease test or two antral biopsies for evaluation of success of antimicrobial therapy for H. pylori infection will result in a false declaration of cure in at least 5% of cases. Three large cup gastric mucosal biopsies for histology are recommended for evaluation of the success of anti-H. pylori therapy.

Bacteriological Techniques↗
PubMed1995

The crystal structure of urease from Klebsiella aerogenes.

The crystal structure of urease from Klebsiella aerogenes has been determined at 2.2 A resolution and refined to an R factor of 18.2 percent. The enzyme contains four structural domains: three with novel folds playing structural roles, and an (alpha beta)8 barrel domain, which contains the bi-nickel center. The two active site nickels are 3.5 A apart. One nickel ion is coordinated by three ligands (with low occupancy of a fourth ligand) and the second is coordinated by five ligands. A carbamylated lysine provides an oxygen ligand to each nickel, explaining why carbon dioxide is required for the activation of urease apoenzyme. The structure is compatible with a catalytic mechanism whereby urea ligates Ni-1 to complete its tetrahedral coordination and a hydroxide ligand of Ni-2 attacks the carbonyl carbon. A surprisingly high structural similarity between the urease catalytic domain and that of the zinc-dependent adenosine deaminase reveals a remarkable example of active site divergence.

Binding Sites↗
PubMed1994

A possible urease-based homologous ELISA for buffalo prolactin.

The feasibility of a direct binding as well as competitive ELISA using urease labelled probe is described for buffalo pituitary prolactin. Rabbit anti-buPRL serum and goat anti-rabbit IgG conjugated to urease were used as primary and secondary antibodies respectively. Assay was performed in polystyrene wells pretreated with 3-amino-propyltriethoxysilane (APTES) compound. APTES pretreatment provided a more receptive surface for prolactin (PRL). In direct binding mode the assay could detect 24 pg of PRL per well. In competition mode the detection limit was less than 0.1 ng of competitor PRL per well. Urease-based ELISA was found highly sensitive when the substrate solution containing pH indicator, bromocresol purple, was used at lower pH (4-4.2). No false positive reactions occurred in the control wells. A good parallelism between buffalo serum sample inhibition curve and the standard PRL inhibition curve was observed.

Animals↗
PubMed1993

Comparison of the urease test and of direct smear examination in the control of treatment of Helicobacter pylori-induced infection.

The sensitivity and specificity of the preformed urease test and of carbolfuchsin-stained smears for the diagnosis of the presence of Helicobacter pylori in gastric mucosa were evaluated before and after antimicrobial treatment. The results obtained by culture were used as the reference point. We studied 41 patients with endoscopically diagnosed duodenal ulcer. Twenty-five of these were treated with furazolidone (100 mg t.i.d.), amoxicillin (500 mg t.i.d.) and metronidazole (250 mg t.i.d.) for 5 days and then with only furazolidone (100 mg t.i.d.) for an additional 25 days. The 16 control patients were treated with cimetidine (800 mg, 4 times a day). The sensitivity of the urease test and of direct smear examination was 100% before treatment and 84.6% and 92.3%, respectively, after treatment. We conclude that the urease test and carbolfuchsin-stained smears, which are highly sensitive for H. pylori diagnosis, present reduced sensitivity when they are employed for the follow-up of patients treated with antimicrobials.

Adult↗
PubMed1995

Evaluation of the severity of Helicobacter pylori infection with urease test: its correlation with histopathology and bacterial density.

In 69 patients, the severity of Helicobactor pylori (H. pylori) infection was evaluated by bacterial density of tissue implants and inflammatory responses by histology. The specimens were taken from gastric angle and antrum (greater and lesser curvature sides) by gastroduodenal endoscopy. In urease test, the severity was measured in 3 grades according to color change of the agar: those change are within 30 minutes (grade 3), 30 minutes to 3 hours (grade 2), and 3 to 6 hours (grade 1), respectively; while the grade 0 indicated no color change occurring 6 hours after tissue inoculation. The severity of infection was assessed according to the bacterial density under high power microscopic fields (Gram's stain). Grade 0 indicated no bacterium seen; grade 1, only 1 to 10 bacteria at all fields; grade 2, 1 to 3 bacteria in each high power field; and grade 3 was 4 bacteria or more on average in each high power field. The degree of inflammatory response was evaluated by inflammatory cell infiltration (H & E stain) and classified into grade 0, 1 and 2, which indicated the inflammatory cell infiltration below 50%, between 50% and 75%, and above 75%, respectively. There are no positive relationships among urease test reaction time, bacterial density grading and degrees of inflammatory cell infiltration. Clinically, the reaction time of urease test cannot reflect the severity of H. pylori infection semi-quantitatively, either in terms of bacterial density or cellular inflammatory response.

Adult↗
PubMed1977

Inhibition of urease by miscellaneous ions and compounds. Implications for the therapy of infection-induced urolithiasis.

One hundred forty-eight drugs and other organic and inorganic substances were screened for their ability to inhibit the enzyme urease in an in vitro system modeled on infected urine. The reported urease-inhibiting properties of ascorbic acid, tetracyclines, and sulfanilamide were not confirmed. At least 50 per cent inhibition was observed in the presence of kanamvcin, hydroxguanidine, benzoquinone, 1,2-naphthaquinone-4-sulfonate, chloramine-T, N-bromoacetamide, copper, mercury, and fluoride. It is, however, unlikely that therapeutically effective concentrations can be attained in urine without giving dosages likely to result in toxic effects. Hydroxyurea, at the dose level used in cytotoxic therapy, may be expected to produce effective inhibition of bacterial urease in the urinary tract, providing renal function is unimpaired and providing urinary volume does not exceed 1 liter per 24 hr. Acetohydroxamic acid is potentially the most useful drug for the treatment of infection-induced urinary stone disease available at present.

Bacterial Infections↗
PubMed1996

The accuracy of the rapid urease test and 13C-urea breath test in the diagnosis of Helicobacter pylori infection.

BACKGROUND: Helicobacter pylori (H. pylori) is an important predisposing factor in peptic ulcer disease. Many tests have been proposed, but there is no generally accepted single method for the detection of H. pylori. This study compared the four available methods in the detection of H. pylori. METHODS: One hundred and thirteen patients were studied with endoscopic biopsy. Biopsy specimens were examined with modified Giemsa stain and rapid urease (CLO) test. Serology (ELISA) and 13C-urea breath test (13C-UBT) were also performed. The 13C-UBT results were expressed at an excess delta 13CO2 excretion of 5 per mil as the upper limit. Multiple breath samples were collected 15, 30 and 60 minutes following 13C-urea ingestion (t = 15, 30, 60) in the first 60 patients. Gastric inflammatory changes were graded according to the Whitehead classification. The diagnostic gold standard was defined when three or more of the four test parameters showed positive. RESULTS: According to this diagnostic gold standard, the positive rates of H. pylori were 97.9% for duodenal ulcer, 81.8% for gastric ulcer, 47.6% for symptomatic gastritis and 13.6% for asymptomatics. Rapid urease test and the 13C-UBT had better sensitivity (93.6% and 96.2%) and accuracy (93.8% and 93.8%). The specificity and positive predictive value for rapid urease test was better than 13C-UBT (94.3% v.s. 88.6%, 97.3% v.s. 94.9% respectively). Modified Giemsa stain had the lowest sensitivity (87.2%), and the ELISA test had the lowest specificity (71.4%). Severity of the gastric inflammatory processes was directly correlated with the excess delta 13CO2 (r = 0.576). CONCLUSIONS: Both the CLO and 13C-UBT had higher accuracy in the detection of H. pylori. When the CLO test result is positive, there is little additional diagnostic benefit from performing other tests. If patients refuse endoscopic examination, 13C-UBT is a good alternative for the detection of H. pylori, either during diagnosis or follow-up after therapy.

Adult↗
PubMed1995

A comparative study between rapid urease (modified), CLO test, culture and histopathological examination for Helicobacter pylori in patients with acid peptic diseases.

A modified Rapid urease test developed by us was evaluated as a screening test for Helicobacter pylori (H. pylori) during and endoscopy survey on patients with Acid Peptic Diseases (APD) and Non Ulcer Dyspepsia (NUD). This was compared with commercially available CLO (Campylobacter Like Organism) test, culture and histopathological examination. The modified Rapid urease test gave a sensitivity of 89.83% and a specificity of 100%, when compared to 95% sensitivity and specificity for commercially available CLO test. Our modified Rapid urease test is simple, economical and a quick test in identifying H. pylori in routine screening of patients with APD and NUD.

Adolescent↗
PubMed1998

Helicobacter pylori and plasma ammonia levels in cirrhotics: role of urease inhibition by acetohydroxamic acid.

BACKGROUND AND AIMS: The role of Helicobacter pylori as a cause of hyperammonaemia in cirrhotics has still not been fully clarified. This study was aimed at evaluating the effect of acute Helicobacter pylori urease inhibition by oral acetohydroxamic acid administration on blood ammonia levels in cirrhotic patients. METHODS: Twenty-nine cirrhotics (14 males, 15 females; mean age: 63 years; Child-Pugh class: 14 A, 9 B, and 6 C) undergoing upper gastrointestinal endoscopy were enrolled in the study. The presence of Helicobacter pylori infection was assessed by rapid urease test and histology. A semi-quantitative grading of bacterial density was also performed at histology. All patients received oral acetohydroxamic acid 750 mg, and blood samples for assessment of ammonia levels were taken before and at 15, 30, 60 and 90 minutes after administration. RESULTS: Helicobacter pylori infection was detected in 20 patients, while 9 patients were uninfected. Acetohydroxamic acid administration led to a significant reduction in blood ammonia levels at 15 and 30 minutes (mean +/- SD, 113 +/- 44 vs 101 +/- 43 and 93 +/- 38 micrograms/dl, respectively; p = 0.002) only in patients with Helicobacter pylori infection. Moreover, the reduction was statistically significant only in Child-Pugh B/C class patients and in those with moderate/marked Helicobacter pylori density in gastric mucosa. Basal ammonia levels did not differ between Helicobacter pylori positive and negative patients, nor in patients with mild and moderate/marked Helicobacter pylori density in gastric mucosa, while Child-Pugh class B/C cirrhotics had higher basal ammonia levels than class A cirrhotics, in both Helicobacter pylori positive and negative groups. CONCLUSIONS: Our data showed that Helicobacter pylori urease inhibition by acetohydroxamic acid administration significantly reduces blood ammonia levels in patients with more advanced liver cirrhosis and in those with a high bacterial density in gastric mucosa.

Aged↗
PubMed1996

Dinuclear Nickel(II) Complexes as Models for the Active Site of Urease.

Dinuclear nickel(II) complexes of the ligands 2,6-bis[bis((2-benzimidazolylmethyl)amino)methyl]-p-cresol (bbapOH), N,N,N',N'-tetrakis(2-benzimidazolylmethyl)-2-hydroxy-1,3-diaminopropane (tbpOH), N-methyl-N,N',N'-tris(2-benzimidazolylmethyl)-2-hydroxy-1,3-diaminopropane (m-tbpOH) and 1-[N,N-bis(2-benzimidazolylmethyl)amino]-3-[2-(3,5-dimethyl-1H-pyrazol-1-yl)ethoxy]-2-hydroxypropane (bpepOH) were prepared in order to model the active site of urease. The novel asymmetric structures of the dinuclear complexes were characterized by X-ray structure analysis. The complex [Ni(2)(bbapO)(ClO(4))(H(2)O)(MeOH)](ClO(4))(2).Et(2)O, 1, crystallizes in the monoclinic space group P2(1)/c, with a = 10.258(2) Å, b = 19.876(3) Å, c = 25.592(4) Å, and beta = 97.12(2) degrees. The nickel ions in 1 are bridged by the phenoxy donor of the ligand and a perchlorate anion. The complexes [Ni(2)(tbpO)(MeCOO)(H(2)O)](ClO(4))(2).H(2)O.Et(2)O, 2, [Ni(2)(m-tbpO)(PhCOO)(EtOH)(2)](ClO(4))(2).EtOH, 3, and [Ni(2)(bpepO)(MeCOO)(H(2)O)(2)](ClO(4))(2).H(2)O.Et(2)O.2EtOH, 4, also crystallize in the monoclinic crystal system with the following unit cell parameters: 2, C2/c, a = 35.360(13) Å, b = 10.958(3) Å, c = 24.821(10) Å, beta = 103.55(3) degrees; 3, Cc, a = 14.663(5) Å, b = 32.630(13) Å, c = 9.839(3) Å, beta = 92.49(2) degrees; 4, C2/c, a = 27.689(13) Å, b = 12.187(5) Å, c = 31.513(14) Å, beta = 115.01(3) degrees. The dinuclear centers of all these complexes are bridged by the alkoxy donor of the ligand and a carboxylate function. Compounds 2 and 3 have one of the nickel ions in a five-coordinated, trigonal bipyramidal coordination environment and thus show a high structural similarity to the dinuclear active site of urease from Klebsiella aerogenes. Furthermore, their magnetic and spectroscopic properties were determined and related to those of the urease enzymes. Activity toward hydrolysis of test substrates (4-nitrophenyl)urea, 4-nitroacetanilide, 4-nitrophenyl phosphate or bis(4-nitrophenyl) phosphate by the dinuclear complexes were examined by UV spectroscopic measurements.

Journal Article↗
PubMed1989

Evidence that the adverse effect of urea fertilizer on seed germination in soil is due to ammonia formed through hydrolysis of urea by soil urease.

Studies using seeds of wheat (Triticum aestivum L.), rye (Secale cereale L.), barley (Hordeum vulgare L.), and corn (Zea mays L.) indicated that the adverse effect of urea fertilizer on seed germination in soil is due to ammonia formed through hydrolysis of urea by soil urease and is not due to urea itself, to urea fertilizer impurities such as biuret, or to nitrite formed by nitrification of urea nitrogen. Support for this conclusion was obtained from (i) comparison of the effects on seed germination in soil of purified urea, urea fertilizers, urea fertilizer impurities, and compounds formed by enzymatic and microbial transformations of urea in soil; (ii) studies showing that ammonia volatilized from soils treated with urea completely inhibited germination of seeds close to, but not in contact with, these soils; and (iii) experiments showing that the adverse effect of urea fertilizer on seed germination in soil was completely eliminated when the soil was autoclaved to destroy urease or was treated with phenylphosphorodiamidate to inhibit soil urease activity before treatment with urea fertilizer.

Journal Article↗
PubMed1977

Nitrogen Metabolism in Soybean Tissue Culture: II. Urea Utilization and Urease Synthesis Require Ni.

Potassium citrate (10 mM, pH 6) inhibits the growth of cultured (Glycine max L.) cells when urea is the sole nitrogen source. Ureadependent citrate toxicity is overcome by three separate additions to the growth medium: (a) NH(4)Cl (20 mM); (b) high levels of MgCl(2) (10 mM) or CaCl(2) (5-10 mM); (c) low levels of NiSO(4) (10(-2) mM). Additions of 10(-2) mM NiSO(4) not only overcome citrate growth inhibition but the resultant growth is usually better than urea-supported growth in basal medium (neither added citrate nor added nickel). In the absence of added citrate, exceedingly low levels of NiSO(4) (10(-4) mM) strongly stimulate urea-supported growth in suspension cultures.Citrate does not inhibit growth when arginine is sole nitrogen source. However, cells using arginine have no net urease synthesis in the presence of 10 mM potassium citrate. When 10(-2) mM NiSO(4) is added to this medium, urease specific activity is 10 times that observed in basal medium lacking both citrate and added nickel.Citrate is a chelator of divalent cations. That additional Mg(2+) or Ca(2+) alleviates urea-dependent citrate toxicity indicates that citrate is acting by chelation, probably of another trace divalent cation; this is probably Ni(2+) since at 10(-2) mM it overcomes citrate toxicity and at 10(-4) mM it stimulates urea-supported growth in the absence of citrate. That ammonia overcomes citrate toxicity indicates that the trace Ni(2+) is essential specifically for the conversion of urea to ammonia. Ni(2+) stimulation of urease levels in arginine-grown cells supports this contention.In basal medium, soybean cells grow slowly with urea nitrogen source presumably because the trace amounts of Ni(2+) present (</=10(-6) mM) are growth-limiting.

Journal Article↗
PubMed2005

First report on autochthonous urease-positive Trichophyton rubrum (T. raubitschekii) from South-east Europe.

BACKGROUND: Trichophyton raubitschekii is a dermatophyte belonging to the T. rubrum complex and is differentiated principally by its positive urease activity and production of profuse macroconidia and microconidia in culture. It is classically isolated from African, South-east Asian and Australian aboriginal patients with tinea corporis or tinea cruris. OBJECTIVES: This study was undertaken to screen Greek and Bulgarian clinical isolates identified as T. rubrum for T. raubitschekii and to delineate these strains by two molecular methods used for the first time in T. rubrum epidemiological studies. METHODS: Ninety-five Greek and 10 Bulgarian strains, originating from various body sites, initially identified as T. rubrum, were screened for urease activity. The biochemical properties and morphology of the urease-positive strains were determined. Strains were delineated with polymerase chain reaction (PCR)-ribotyping amplifying repeat elements of the intergenic spacer region and by PCR fingerprinting. RESULTS: Five Greek and one Bulgarian T. raubitschekii strains were identified comprising isolates from patients with tinea manuum (one), tinea corporis (one), tinea cruris (one) and tinea unguium (three). Only one strain had the classical T. raubitschekii microscopic morphology, whereas the remaining five presented a dominant arthroconidial phenotype. Both typing methods clustered all T. raubitschekii and T. rubrum isolates together in the same group, indicating strain homogeneity in the genetic regions examined. CONCLUSIONS: The reported isolation of T. raubitschekii in the Balkan and South-eastern Mediterranean regions extends the geographical distribution of this species. As the more primitive T. raubitschekii probably represents the parental population of T. rubrum, the Greek and Bulgarian T. raubitschekii strains could represent a remnant of the T. rubrum spread that took place after the First World War, rather than being a recent epidemiological event.

Bulgaria↗
PubMed1994

The effects of sodium citrate and oral potassium citrate on urease-induced crystallization.

OBJECTIVES: To study the effects of citrate on urease-induced crystallization in human urine. MATERIALS AND METHODS: Urine samples were collected from seven healthy volunteers with no history of urinary tract infection or stone disease. Citrate was removed from the urine samples by decomposition with citrate lyase. Citrate was then added to the urine in increasing concentrations. Oral potassium citrate was given to the volunteers and their urine was collected. The samples were incubated with urease and the crystallization induced was observed by the Coulter counter technique, by using an optical microscope and by precipitated material analysis. RESULTS: The initiation of crystallization was markedly delayed by both the addition of citrate to the urine and after the ingestion of citrate. Crystal growth and the resulting precipitation of both calcium and magnesium showed a concentration-dependent reduction when citrate was added up to a concentration of 4 mM. Crystal growth and precipitation of calcium and magnesium were also significantly decreased by oral citrate intake. CONCLUSION: Citrate added to the urine or taken orally markedly delays urease-induced crystallization in human urine.

Administration, Oral↗
PubMed1991

Intralysosomal generation of ammonia from urea by endocytosed urease results in secretion of free lysosomal arylsulfatase-A and increased activity of membrane-bound beta-glucosidase in cultured brain cells.

Hyperammonemia interferes with normal brain function. The effect of ammonia on free and membrane-bound lysosomal enzymes and on mucopolysaccharide metabolism was studied in cultured rat brain cells (ROC-1, hybridoma between C6-astrocytoma and oligodendrocytes). Intralysosomal ammoniagenesis was achieved from urea by endocytosed Jackbean urease followed by incubation of the cultures with urea. The intralysosomal location of urease was evidenced by the protective effects of leupeptin and urea on the stability of intracellular urease. Ammonia formed from urea resulted in an increased secretion of lysosomal arylsulfatase-A (AS-A), but not of the membrane-bound lysosomal beta-glucosidase into the culture medium, thus intralysosomal AS-A activity decreased. Lysosomal, membrane-bound beta-glucosidase activity increased, presumably due to intralysosomal proteolytic protection following an increased lysosomal pH. Intralysosomal ammoniagenesis temporarily impaired 35SO4-glycosaminoglycan degradation of prelabeled cells. The results support the hypothesis that hyperammonemic states may interfere with lysosomal functions in vivo as well in cultured cells.

Ammonia↗