Biomineralization of soil with crude soybean urease using different calcium salts

Calcium salt is an important contributing factor for calcium-based biomineralization.To study the effect of calcium salt on soil biomineralization using crude soybean urease,the calcium salts,including the calcium chloride (CaCl_(2)),calcium acetate ((CH_(3)COO)_(2)Ca) and calcium nitrate (Ca(NO_(3))_(2)),were used to prepare the biotreatment solution to carry out the biomineralization tests in this paper.Two series of biomineralization tests in solution and sand column,respectively,were conducted.Scanning electron microscopy (SEM) and X-ray diffraction (XRD) were performed to determine the microscopic characteristics of the precipitated calcium carbonate (CaCO_(3)) crystals.The experimental results indicate that the biomineralization effect is the best for the CaCl2 case,followed by (CH_(3)COO)_(2)Ca,and worst for Ca(NO_(3))_(2) under the test conditions of this study (i.e.1 mol/L of calcium salt-urea).The mechanism for the effect of the calcium salt on the biomineralization of crude soybean urease mainly involves: (1) inhibition of urease activity,and (2) influence on the crystal size and morphology of CaCO_(3).Besides Ca^(2+) ,the anions in solution can inhibit the activity of crude soybean urease,and NO_(3)− has a stronger inhibitory effect on the urease activity compared with both CH_(3)COO^(−) and Cl^(−) .The co-inhibition of Ca^(2+) and NO_(3)− on the activity of urease is the key reason for the worst biomineralization of the Ca(NO_(3))_(2) case in this study.The difference in biomineralization between the CaCl_(2) and (CH_(3)COO)_(2) Ca cases is strongly correlated with the crystal morphology of the precipitated CaCO_(3).

Activities of Urease and Phosphatase in Integrated Vertical Flow Constructed Wetland and Purification Effect of Black and Malodorous River

[Objective]The activities of urease and phosphatase in integrated vertical flow constructed wetland and purification effect of black and malodorous river were studied.[Method]Honeycomb stone and soft filler were used as matrix,the biological grid integrated vertical flow constructed wetland system was built to strengthen treatment for black and malodorous river,and the activities of urease and phosphatase of two fillers in different functional areas of constructed wetland were tested to study the film formation property of two fillers in different functional areas.[Result]Both urease and phosphatase activities on biofilm of soft filler were significantly higher than on biofilm of honeycomb stone filler;indicating that choosing soft filler was more advantageous to the growth of biofilm;and urease activity had significant correlation with total nitrogen removal rate,while phosphatase activity had no significant correlation with total phosphorus removal rate.[Conclusion]This study provided evidences for utilizing the activities of urease and phosphatase to evaluate the purification effect of black and malodorous river and choose appropriate filler.

Dynamic Effects of PAEs on Soil Urease and Phosphatase

[Object] The study aimed to supply a reference for evaluating ecotoxicology of soil contaminated with phthalate acid esters(PAEs).[Method] The dynamic effects of DBP and DEHP on activities and kinetics parameters of urease and phosphatase in agro-soil contaminated artificially with DBP and DEHP were studied.[Result] The activities of urease and phosphatase were both inhibited significantly by higher contents of DBP and DEHP in soils compared with CK.The inhabitations increased with increasing DBP and DEHP c...

Single and joint effects of pesticides and mercury on soil urease

The influence of two pesticides including chlorimuron-ethyl and furadan and mercury （Hg） on urease activity in 4 soils （meadow burozem and phaeozem） was investigated. The soils were exposed to various concentrations of the two pesticides and Hg individually and simultaneously. Results showed that there was a close relationship between urease activity and organic matter content in soil. Chlorimuron-ethyl and furadan could both activate urease in the 4 soils. The maximum increment of urease activity by chlorimuronethyl was up to 14%-18%. There was almost an equal increase （up to 13%-21%） in the urease activity by furadan. On the contrary, Hg markedly inhibited soil urease activity. A logarithmic equation was used to describe the relationship （P〈0.05） between the concentration of Hg and the activity of soil urease in the 4 tested soils. Semi-effect dose （ED50） values by the stress of Hg based on the inhibition of soil urease in the 4 soils were 88, 5.5, 24 and 20 mg/kg, respectively, according to the calculation of the corresponding equations. The interactive effect of chlorimuron-ethyl or furadan with metal Hg on soil urease was mainly synergic at the highest tested concentrations.

Effect of Combined Heavy Metal Pollution on Nitrogen Mineralization Potential,Urease and Phosphatase Activities in a Typic Udic Ferrisol

Individual and combined effects of Cu, Pb, Zn and Cd on N mineralization, urease and phosphatase were examined in a Typic Udic Ferrisol in laboratory by employing an uniform design and a single factor design.Soil pollution caused by heavy metals inhibited N mineralization (No value) and urease and phosphatase activities. The combined pollution of metals alleviated their toxicity to N mineralization to some extent,whereas aggravated the toxicity to urease and phosphatase. Phosphorous application could mitigate the toxic effect of heavy metals on phosphatase activities, while alleviating effect of N application on the toxicity of heavy metals to urease was inconsistent. However, the mitigating effect of the fertilizers was limited in heavily polluted soils.

Dual specimens increase the diagnostic accuracy and reduce the reaction duration of rapid urease test

AIM:To evaluate the influence of multiple samplings during esophagogastr oduodenoscopy(EGD) on the accuracy of the rapid urease test,and the validity of newly developed rapid urease tests,HelicotecUT plus test and HelicotecUT test,CLO test and ProntoDry test.METHODS:A total of 355 patients undergoing EGD for dyspepsia were included.Their Helicobacter pylori(H.pylori) treatment status was either nave or eradicated.Six biopsy specimens from antrum and gastric body,respectively,were obtained during EGD.Single antral specimens and dual(antrum+body) specimens were compared.Infection status of H.pylori was evaluated by three different tests:culture,histology,and four different commercially available rapid urease tests(RUTs)-including the newly developed HelicotecUT plus test and HelicotecUT test,and established CLO test and ProntoDry test.H.pylori status was defined as positive when the culture was positive or if there were concordant positive results among histology,CLO test and ProntoDry test.RESULTS:When dual specimens were applied,sensitivity was enhanced and RUT reaction time was signif icantly reduced,regardless of their treatment status.Thirty minutes were enough to achieve an agreeable positive rate in all the RUTs.Both newly developed RUTs showed comparable sensitivity,specif icity and accuracy to the established RUTs,regardless of patient treatment status,RUT reaction duration,and EGD biopsy sites.CONCLUSION:Combination of antrum and body biopsy specimens greatly enhances the sensitivity of rapid urease test and reduces the reaction duration to 30 min.

Oral Immunization of Mice With Vaccine of Attenuated Salmonella typhimurium Expressing Helicobacter pylori Urease B Subunit

Objective To prepare the live recombinant vaccine of attenuated Salmonella typhimurium SL3261 expressing Helicobacterpylori （H. pylori） B subunit （UreB） and to determine whether it could be used as an oral vaccine against H. pylori infection. Methods Using genomic DNA of H. pylori Sydney strain （SS1） as template, the H. pylori UreB gene fragment was amplified by PCR and subcloned into the expression vector pTC01. The recombinant plasmid pTC01-UreB was then transferred into LBS000 to obtain modified forms, and further conversed into the attenuated Salmonella typhimurium SL3261 to obtain recombinant SL3261/pCT01-UreB as an oral immunization reagent, which was then used to orally immunize Balb/c mice twice at a three-week interval. Twelve weeks later, anti-UreB IgA antibodies in intestinal fluid and IgG antibodies in sera were determined by ELISA. The relating data in control groups （including body weight, gastric inflammation, etc.） were also collected. Results The sequencing analysis showed that the UreB gene fragment amplified by PCR was consistent with the sequence of the H. pylori UreB gene. The restriction enzyme digestion revealed that the correct pTC01-UreB was obtained. SDS-PAGE and Western blot showed that a 61KD protein was expressed in SL3261/pTC01-UreB, which could be recognized by anti-H, pylori UreB antiserum and was absent in the control containing only Salmonella typhimurium SL3261 strain. The multiple oral immunization with SL3261/pTC01-UreB could significantly induce H. pylori specific mucosal IgA response as well as serum IgG responses. IFN-T and IL-10 levels were significantly increased in SL3261/pTC01-UreB group, and no obvious side effect and change in gastric inflammation were observed. Conclusion The attenuated vaccine of Salmonella typhimurium expressing H. pylori UreB can be used as an oral vaccine against H. pylori infection.

Preparation of Glutaraldehyde Cross-Linked Chitosan Beads Under Microwave Irradiation and Properties of Urease Immobilized onto the Beads

The glutaraldehyde cross-linked chitosan beads were prepared under microwave irradiation and urease was immobilized onto the beads. The activity and the yield of enzyme activity of the immobilized urease were 10.83 U/g carrier and 47.700, respectively. The optimum conditions of immobilization were 100 of glutaraldehyde volume fraction, 10 mg/g of urease/beads weight ratio, 24 h of the processing time and pH 6.5 of the reaction medium for immobilization. The properties of the immobilized urease were investigated and compared with those of the free enzyme. The optimum pH values were 6.5 and 7.0 for the immobilized and free urease, respectively. The optimum temperature was 60 ℃ for the free urease, while it shifted to 65 ℃ for the immobilized enzyme. The Michaelis constant Km was 9.1 mmol/L for the immobilized and 12.5 mmol/L for the free urease. The immobilized urease retained 4000 of its initial enzyme activity even after 10 repeated uses. The immobilized urease stored at 4 ℃ retained 4600 of its initial activity even after 35 d.

Evaluating the immune responses of mice to subcutaneous immunization with Helicobacter pylori urease B subunit

Background： Helicobacter pylori, a gram-negative bacterial pathogen that expresses a strong urease activity, is associated with the development of gastroduodenal disease. Urease B subunit, one of the two structural subunits of urease, was expressed in E. coil BL21 （DE3） strain. The objective of this study was to evaluate the effects of He/icobacter py/ori urease B subunit on the immune responses in mice by subcutaneous immunization. Methods： The mice were immunized and boosted with Helicobacter pylori urease B subunit antigen subcutaneously three times with 2-wk intervals between the immunizations and boosters. The mice in the control group were immunized with PBS. The adjuvant group received PBS containing complete/incomplete freund＇s adjuvant identical to antigen group without Helicobocter pylori urease B subunit antigen. Four weeks after the final booster, all the mice were sacrificed. Blood was collected on d 0, 14, 28 and 56 before immunization, booster and sacrifice, respectively. Immediately after sacrifice, gastric liquid and spleen were collected for antibody and cytokine analyses. Results： Urease B subunit increased the concentrations of serum and gastric anti-urease B antigen specific IgG, and the levels of interteukin-4 and interferon-y in splenocytes of the mice （P 〈 0.05）. Conclusions： This study demonstrated that recombinant responses in mice by subcutaneous immunization, which against Helicobocter pylori. urease B subunit can induce systemic and local immune might be used as the effective component of vaccine

Characteristics of Soybean Urease Mineralized Calcium Carbonate and Repair of Concrete Surface Damage

The C60 concrete blocks with surface crack damage under high temperature environment were soaked by adding appropriate amount of soybean urease into the CO(NH_(2))_(2)-CaCl_(2) solution,the soybean urease mineralized calcium carbonate were characterized,and the effect of repairing concrete surface crack damage were evaluated by the surface sedimentation of C60 concrete blocks in the study.The experimental results showed that the activity of soybean powder was statistically significant,and its productivity of urease was comparable with that of urease-producing bacteria.After immersion in a soybean solution,a layer of complete and continuous white sediment covered the concrete surface.The cracks on the concrete surface were completely shielded,and the rising temperature on infrared thermal image of the concrete after repair was lower than before.Besides,through analysis by SEM,EDS,and XRD,the products formed after repair were found to be calcite-type CaCO_(3) with high purity,and the crystals exhibited different morphological features.The above results indicate that soybean urease can regulate and induce the formation of calcium carbonate,and the precipitate is innocuous and harmless,suitable for a new type of concrete crack repair material.

Response of Nematodes in a Hapli-Udic Argosol to Urea Amended with Urease and Nitrification Inhibitors

Effects of urea amended with urease and nitrification inhibitors on soil nematode communities were studied in a Hapli- Udic Argosol （Cambisol, FAO） in Liaoning Province of Northeast China. A completely random design with four treatments, i.e., conventional urea （CU）, slow-release urea amended with a liquid urease inhibitor （SRU1）, SRU1 ＋nitrification inhibitor dicyandiamide （SRU2）, and SRU1 ＋ nitrification inhibitor 3,5-dimethylpyrazole （SRU3） and four replicates were applied. Thirty-nine genera of nematodes were identified, with Cephalobus and Aphelenchus being dominant; and in all treatments, the dominant trophic group was bacterivores. In addition, during the growth period of spring wheat （Triticum aestivum L.）, soil urease activity was lower in SRUs than in CU. The numbers of total nematodes and bacterivores at wheat heading and ripening stages, and omnivores-predators at ripening stage were higher in SUR3 than in CU, SRU1 and SRU2 （P 〈 0.05）.

Soil Urease Activity of Sundarban Mangrove Ecosystem, India

Vertical occurrence of soil urease activity along with ammonia content from three distinct regions viz. Deep forest region (No tidal action and wave attack occurs as it is furthest from river shore and it contains maximum content of organic carbon and minimum soil salinity and silicate concentration. In this zone plenty of pneumatophores, below ground root and dense vegetation are found), Rooted region (It is situated in between Deep forest region and Un-rooted region. This region contains only pneumatophores but it is devoid of long roots and vegetations. It faces wave attack and tidal action less than that of Un-rooted region) and Un-rooted region (It is closest to river shore and faces maximum wave attack and tidal action;it contains minimum organic carbon but maximum soil salinity and silicate concentration. This zone is totally devoid of any roots, pneumatophores and vegetations) of Sundarban mangrove forest ecosystem, India revealed an interesting explanation. Soil urease activity showed a decreasing pattern with increase in depth from the deep forest region of the Sundarban forest ecosystem. Soil urease activity was found to be more sensitive to soil temperature and pH rather than soil salinity. This ensured that soil urease along with the microbes present in the Sundarban forest ecosystem are more tolerant to fluctuation in salinity than that of temperature. Soil ammonia concentration was found to be directly governed by the soil urease activity [The regression equation is Ammonia in soil = -1.64 + 0.0402 Urease Activity (R-Sq = 62.9%, P < 0.001, n = 41)].

Nickel trafficking system responsible for urease maturation inHelicobacter pylori

Helicobacter pylori(H.pylori)is a common human pathogen responsible for various gastric diseases.This bacterium relies on the production of urease and hydrogenase to inhabit the acidic environment of the stomach.Nickel is an essential cofactor for urease and hydrogenase.H.pylori has to uptake sufficient nickel ions for the maturation of urease,and on the other way,to prevent the toxic effects of excessive nickel ions.Therefore,H.pylori has to strike a delicate balance between the import of nickel ions,its efficient intracellular storage,and delivery to nickel-dependent metalloenzymes when required.The assembly and maturation of the urease enzyme is a complex and timely ordered process,requiring various regulatory,uptake,chaperone and accessory proteins.In this review,we focus on several nickel trafficking proteins involved in urease maturation:NikR,NixA,HypAB,UreEFGH,HspA,Hpn and Hpnl.The work will deepen our understanding of how this pathogenic bacterium adapts to severe habitant environments in the host.

Detection of Helicobacter pylori:A faster urease test can save resources

AIM: To investigate whether differences in the rapidity of a positive result for Helicobacter pylori can save res ources, by comparing two commercially available urease kits. METHODS: One hundred and eighty-five adults (130 outpatients, 55 inpatients) undergoing gastroscopy were entered prospectively. Patients were divided into two groups: Group 1 (if they were not on PPIs, antibiotics, H2A, bismuth or sucralfate for up to 14 d prior to the endoscopy) and Group 2 (if they were on, or had been on, any of the above medication in the previous 14 d). At endoscopy two sets of biopsies, taken in random order, were placed in the wells of the Campylobacter-like organism (CLO) test (Kimberly-Clark, Utah, USA) and the Quick test (Biohit Plc, Helsinki, Finland). Five additional gastric biopsies were taken for histology/Giemsa and immunohistochemical study. The two urease test slides were read at 2 min, 30 min, 2 h and 24 h. Sensitivity and specif icity at 24 h were determined. RESULTS: At 24 h, for all patients, there was no difference in sensitivity (100% vs 97.5%), specificity (99.3%), positive (97.5%) and negative predictive values (100% vs 99.3%) between the CLO and Quick tests, respectively. There was a positive result at 30 min in 17/41 (41.5%) CLO tests, and in 28/40 (70%) Quick tests, P = 0.05. Quick test enabled the prescription of eradication therapy before discharge in all 28/40 patients. Only 12 (30%) follow-up appointments were needed. If the CLO test had been used alone, only 17 (41.5%) prescriptions would have been possible prior to discharge and 24 (58%) follow-up appointments would be needed (P = 0.001). Of 2000 gastroscopies performed annually at our unit, a saving of 123 follow-up appointments (total: 8856 Euros or 11 808 USD) would be achieved if we switched to the Quick test. CONCLUSION: Direct comparison of locally available urease test kits is worthwhile, since the appropriate choice results in a significant saving of resources. Local costs and follow-up protocols will determine the magnitude of these savings.

Experimental Study on Urease Activity and Cementation Characteristics of Soybean

A new and more ecologically sound cementing material known as“bio-cement”has been found to have the capacity to consolidate loose gravel into sand columns offering a certain degree of strength,and to fill and repair cracks in concrete to restore resilience.The typical representative is the microbial induced calcium carbonate deposition technology(MICP)and enzyme induced calcite precipitation(EICP).As part of this research,EICP with soybean urease as the core was studied.The test results show that soybean urease activity is significantly affected by pH and urea concentration values,while the external nickel source is not found to impair a stimulating effect on activity.When the concrete specimens were immersed in the composite solution of soybean urease,urea,and calcium chloride after having been subjected to a high temperature,a continuous layer of white precipitations quickly appeared on the surface of the specimens.Measured using a metalloscope,the thickness of the precipitations was found to reach up to 2.0 mm,while the surface water absorption rate was reduced by 70%.The effects of this combined outcome are believed to significantly protect and improve the durability of the concrete specimens previously subjected to a high temperature.At the same time,the composite solution is shown to be capable of cementing fly ash,with the cubic strength of the finished samples reaching 4.0 MPa after 3 days.Results from the use of a scanning electron microscopy(SEM),energy dispersive spectroscopy(EDS),and X-ray diffraction(XRD),reveal that both the white precipitations on the surface of the concrete specimens and the cement binding the fly ash particles are calcite crystals.It is concluded from these preliminary study results that the use of soybean urease as a bio-cement had proved successful.

Immobilization of urease in metal–organic frameworks via biomimetic mineralization and its application in urea degradation

Enzyme immobilization has been accepted as an efficient technique for improving the stability and recyclability of enzymes.Herein,biomimetic mineralization strategy was employed to achieve the immobilization of urease in a type of metal–organic frameworks(zeolite imidazolate framework-8,ZIF-8),and the immobilized enzyme urease@ZIF-8 was systematically evaluated for its structure,activity,stability and recyclability,using the hydrolysis of urea as a model.The entrapment of urease was found to be realized in a synchronous manner with the formation of ZIF-8 crystal.The loading of urease in ZIF-8 was measured to be ca.10.6%through the bicinchoninic acid(BCA)protein assay.The encapsulated urease could efficiently maintain its native conformation,which endowed the immobilized urease with excellent activity and stability,even in harsh conditions(e.g.,in the presence of trypsin,acidic or alkali conditions,or at high temperature).Further,urease@ZIF-8 exhibited good recyclability during the degradation of urea,in which it could keep 58.86%of initial activity after being used for 5 cycles.Thus,biomimetic mineralization could be potentially utilized as a promising method to prepare immobilized ureases with superior activity,stability and recyclability,thereby facilitating the construction of efficient catalysts for industrial biocatalysis and biosensing.

Intraprocedural gastric juice analysis as compared to rapid urease test for real-time detection of Helicobacter pylori

BACKGROUND Endofaster is an innovative technology that can be combined with upper gastrointestinal endoscopy(UGE)to perform gastric juice analysis and real-time detection of Helicobacter pylori(H.pylori).AIM To assess the diagnostic performance of this technology and its impact on the management of H.pylori in the real-life clinical setting.METHODS Patients undergoing routine UGE were prospectively recruited.Biopsies were taken to assess gastric histology according to the updated Sydney system and for rapid urease test(RUT).Gastric juice sampling and analysis was performed using the Endofaster,and the diagnosis of H.pylori was based on real-time ammonium measurements.Histological detection of H.pylori served as the diagnostic gold standard for comparing Endofaster-based H.pylori diagnosis with RUT-based H.pylori detection.RESULTS A total of 198 patients were prospectively enrolled in an H.pylori diagnostic study by Endofasterbased gastric juice analysis(EGJA)during the UGE.Biopsies for RUT and histological assessment were performed on 161 patients(82 men and 79 women,mean age 54.8±19.2 years).H.pylori infection was detected by histology in 47(29.2%)patients.Overall,the sensitivity,specificity,accuracy,positive predictive value,and negative predictive value(NPV)for H.pylori diagnosis by EGJA were 91.5%,93.0%,92.6%,84.3%,and 96.4%,respectively.In patients on treatment with proton pump inhibitors,diagnostic sensitivity was reduced by 27.3%,while specificity and NPV were unaffected.EGJA and RUT were comparable in diagnostic performance and highly concordant in H.pylori detection(κ-value=0.85).CONCLUSION Endofaster allows for rapid and highly accurate detection of H.pylori during gastroscopy.This may guide taking additional biopsies for antibiotic susceptibility testing during the same procedure and then selecting an individually tailored eradication regimen.

Application of a simple calorimetric data analysis on the binding study of cyanide ions by Jack bean urease

Cyanide ion was studied as an effector of Jack bean urease(JBU) at 300 K in 30 mmol/LTris buffer,pH 7 by isothermal titration calorimetry(ITC).The simple novel model was used for CN^- + JBU interaction over the whole range of CN^- concentrations.The binding parameters recovered from the simple novel model were attributed to the cyanide ion interaction.It was found that cyanide ion acted as a noncooperative inhibitor of JBU,and there is a set of 12 identical and independent binding sites for CN^- ions.The di...

Novel Investigation on Ammonium Thiosulphate (ATS) as an Inhibitor of Soil Urease and Nitrification

Among the numerous products so far promoted as effective urease and/or nitrification inhibitors, it is possible to detect a renewed interest in environmentally friendly tools, such as ammonium thiosulphate (ATS, (NH4)2S2O3) which is currently used as fertilizer for N and S nutrition. Among contradictory results accounted in the current literature, there is reported that ammonium thiosulphate (ATS) exerts inhibitory activity at large but unlikely agronomic rates of 2500 - 5000 mg·kg-1 soil. We carried out a novel experiment aimed to: a) verify the inhibitory action of ATS, even when applied in soil at low rates (25 and 100 mg·kg-1 soil as S-ATS), towards urease activity and nitrification in urea treated and not treated soils;b) investigate on the influence of ATS on the soil microbial biomass as it is generally assumed that soil microorganisms are the main agents of urea hydrolysis. For these purposes we selected an arable sandy soil and a grassland sandy-loam soil which are treated with urea or/and ATS. Results obtained from this novel investigation showed that a) ATS significantly decreased urease activity in both soil types and that the exerted inhibitory effect was moderate and short-term;b) ATS retarded the hydrolysis of urea and lowered nitrate production in the urea treated soils;c) the available fraction of iron and manganese in the used soils might be involved in the mechanism of inhibition;d) ATS did not affect the size of soil microbial biomass pool;e) the efficiency of ATS as urease and nitrification inhibitor was more evident in the sandy soil. Because of its properties, ATS may be applicable on many crops without being harmful on the soil microbial pool.

Limitations of urease test in diagnosis of pediatric Helicobacter pylori infection

The diagnosis of Helicobacter pylori(H. pylori) infection is usually based on the results of urease test and histology. The urease test known as a simple and cheap method does not need special skills to perform or to read the result. The time needed for the test to turn positive depends on the concentration of bacteria, and the accuracy is up to the density of H. pylori density in the biopsy sample, which is generally lower in children than adolescents and adults. Therefore, there are debates about the sensitivity of the urease test in children. The reason for lower sensitivity of the urease test in children was not identified, but might be related to the low density and patchy distribution of bacteria. In this review, we discuss the limitations of the urease test in children according to age, histology, number of biopsy samples, and biopsy site. In children under 5 years old, the differences in positivity rate when the urease test used one or three biopsy samples, and samples from the antrum or the gastric body, were larger than those in children aged 5-15 years. Thus, three or more biopsy samples from both the antrum and body would improve the sensitivity of H. pylori infection diagnosis in children under 5 years old.