A low cost rapid urease test to detect Helicobacter pylori infection in resource limited settings.

The aim of the study was to evaluate the suitability of a modified one minute rapid urease test (one day rapid urease test) as a low cost H. pylori detection method. A sample of 205 patients clinically suspected of having H. pylori infection was tested. One day rapid urease test and histology based H. pylori tests (the gold standard) were performed on endoscopic antral biopsies. There were 6 true positives, 191 true negatives, 8 false positives and zero false negatives. The sensitivity, specificity and positive (PPV) and negative predictive values (NPV) of the test were 100%, 96%, 42.9%, and 100% respectively. The cost per patient was 0.3US$. High sensitivity, specificity and NPV, low cost and simplicity of method were the advantages of the test and the main limitation was low PPV. Hence, one day rapid urease test can be considered as a suitable low cost method to detect H. pylori infection in resource limited settings.

In vitro assessment of Helicobacter pylori ureases inhibition by honey fractions.

BACKGROUND AND AIMS: Pathogenic effects associated with Helicobacter pylori infection include peptic ulcer. Pathogenicity is mediated by the bacteria ureases. The emerging resistance of H. pylori to clarithromycin used in combination with amoxicillin in the treatment of H. pylori infection has necessitated the search for other means of combating the scourge of H. pylori, hence the search for potent H. pylori urease inhibitor. The aim of the study was to evaluate honey fractions obtained from different geographic zones for H. pylori urease inhibitory potentials. METHODS: Chloroform and diethyl-ether were used to extract honey fractions (HS) and Manuka honey (HM). H. pylori ureases were obtained from a 48-h culture through sonication. Urease activity was measured spectrophotometrically by assaying the reduction in NADH in coupled urease-glutamate dehydrogenase (GDH) system, whereas urease inhibition was calculated by comparing the NADH oxidation rate before and after incubation with honey fractions. RESULTS: Urease inhibition by the HS and HM were time and concentration independent. Chloroform extract of HS showed 48 and 42% inhibitory activities against urease from H. pylori (369C) and H. pylori (ATCC 43526), respectively, wheeras diethyl ether extract of HM showed 45 and 51% inhibitory activities against urease from H. pylori (369C) and H. pylori (ATCC 43526), respectively. Dixon plot revealed that HM extract showed mixed inhibition pattern in a reversible inhibition kinetics. CONCLUSIONS: Honey fractions were good inhibitors of H. pylori urease and may serve as a template in the development of urease inhibitors in pharmaceuticals.

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 resources, 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 specificity 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.

[Assessment of rate of hydroxyl anions production by Helicobacter pylori in stomach].

Production of hydroxyl anions by tissue samples of pylorus mucous membrane obtained from 45 patients with gastric or duodenal ulcers was investigated. The production was estimated using the recently developed method based on measurement of rate of pH change in urea-containing reaction mixture. The rate of [OH-] generation as a result of H. pylori metabolism accounted on pylorus square varied from 0.4 to 1318.9 mcmol [OH-]/min, and in 90.2% of cases it did not exceed 128.1 mcmol [OH-]/min. This rate is comparable to mean rate of [H+] generation in stomach of healthy man--114.2-238.4 mcmol [H+]/min. Obtained results allow to conclude that this bacterium may participate in regulation of stomach acid-base balance.

Assessment of the two Helicobacter pylori alpha-1,3-fucosyltransferase ortholog genes for the large-scale synthesis of LewisX human milk oligosaccharides by metabolically engineered Escherichia coli.

We previously described a bacterial fermentation process for the in vivo conversion of lactose into fucosylated derivatives of lacto-N-neotetraose Gal(beta1-4)GlcNAc(beta1-3)Gal(beta1-4)Glc (LNnT). The major product obtained was lacto-N-neofucopentaose-V Gal(beta1-4)GlcNAc(beta1-3)Gal(beta1-4)[Fuc(alpha1-3)]Glc, carrying fucose on the glucosyl residue of LNnT. Only a small amount of oligosaccharides fucosylated on N-acetylglucosaminyl residues and thus carrying the LewisX group (Le(X)) was also produced. We report here a fermentation process for the large-scale production of Le(X) oligosaccharides. The two fucosyltransferase genes futA and futB of Helicobacter pylori (strain 26695) were compared in order to optimize fucosylation in vivo. futA was found to provide the best activity on the LNnT acceptor, whereas futB expressed a better Le(X) activity in vitro. Both genes were expressed to produce oligosaccharides in engineered Escherichia coli (E. coli) cells. The fucosylation pattern of the recombinant oligosaccharides was closely correlated with the specificity observed in vitro, FutB favoring the formation of Le(X) carrying oligosaccharides. Lacto-N-neodifucohexaose-II Gal(beta1-4)[Fuc(alpha1-3)]GlcNAc(beta1-3)Gal(beta1-4)[Fuc(alpha1-3)]Glc represented 70% of the total oligosaccharide amount of futA-on-driven fermentation and was produced at a concentration of 1.7 g/L. Fermentation driven by futB led to equal amounts of both lacto-N-neofucopentaose-V and lacto-N-neofucopentaose-II Gal(beta1-4)[Fuc(alpha1-3)]GlcNAc(beta1-3)Gal(beta1-4)Glc, produced at 280 and 260 mg/L, respectively. Unexpectedly, a noticeable proportion (0.5 g/L) of the human milk oligosaccharide 3-fucosyllactose Gal(beta1-4)[Fuc(alpha1-3)]Glc was produced in futA-on-driven fermentation, underlining the activity of fucosyltransferase FutA in E. coli and leading to a reassessment of its activity on lactose. All oligosaccharides produced by the products of both fut genes were natural compounds of human milk.

Evaluation of a liquid urease test (LUT) for detection of Helicobacter pylori.

UNLABELLED: The aim of our study was to develop a rapid diagnostic urease test to demonstrate the presence of Helicobacter pylori in the Endoscopy room. MATERIALS AND METHODS: 200 consecutive patients referred to gastroscopy for different indications, were included in this study. One antral biopsy sample was obtained to be immersed in our test. The same sample was used for histological evaluation, considered to be the gold standard method for diagnose of Helicobacter pylori infection. RESULTS: 135 patients (67.5%) were found positives and 65 patients (32.5%) were negatives in our test. 128 patients (64%) showed Helicobacter pylori on histological examination. Our test showed a sensitivity of 91%, specificity of 88.1%, and positive and negative predictive values of 95% and 80% respectively. A remarkable correlation between density of Helicobacter pylori and reading time was also observed, where a high density of the bacteria reduced the reaction time in this liquid test. Furthermore, an overall accuracy of 90% was shown, which is comparable with other available commercial tests. CONCLUSION: LUT is easy to handle, cost effective and fast, with a high positive predictive value.

Helicobacter pylori culture from a positive, liquid-based urease test for routine clinical use: a cost-effective approach.

BACKGROUND: The aim of our study was to test the feasibility of culturing Helicobacter pylori directly from biopsies aimed for rapid urease test in routine clinical practice. MATERIALS AND METHODS: In 260 consecutive patients referred for gastroscopy because of dyspepsia one antral biopsy was routinely used for our "in house" rapid urease test (RUT). Positive biopsies were placed in a transport medium and sent to the laboratory. The biopsies were cultured and incubated at 37 degrees C for 5-7 days. H. pylori was identified and routinely tested for antimicrobial resistance by using the E-test. RESULTS: In 118 out of 260 patients (45%) the urease test turned positive and the growth of H. pylori was sufficient to allow testing of antimicrobial resistance. CONCLUSION: H. pylori could be cultured from almost all positive RUT specimens. A liquid RUT is thus more suitable for culture, saving additional biopsies.

A home made rapid urease test in the diagnosis of Helicobacter pylori infection.

AIM OF STUDY: To determine whether a homemade rapid urease test (RUT) (1 mL of 10% urea broth in distilled water plus one drop of 1% phenol red as indicator, cost/test USD0.19) was reliable when compared to histology in the diagnosis of HP infection. METHOD: Prospective consecutive sampling of patients who underwent outpatient oesophagogastroduodenoscopy and antral biopsies from October 1996 to January 1997. RUT and histology examinations were done on all specimens. Sensitivity, specificity, positive predictive value, negative predictive value and accuracy of the RUT were calculated accordingly. RESULTS: Amongst our 140 patients, the sensitivity, specificity, positive and negative predictive values and accuracy of RUT were 94%, 99%, 99%, 95% and 96% respectively. Seventy-seven percent of the positive RUTs can be detected within 1 hour. CONCLUSION: Our homemade RUT is an inexpensive test with good sensitivity and specificity for HP infection.

[The appropriate time for the assessment of Helicobacter pylori eradication].

As all of the guidelines on the management of H. pylori infection suggest, the assessment of the eradication is generally performed at least 4 weeks after the completion of eradication treatment. However, H. pylori occasionally re-appears after one month, even the successful eradication was confirmed by the guideline. In this study, we investigated the appropriate time for the assessment of H. pylori eradication, mainly by using 13C urea breath tests (UBT) as a positive standard. From July 1992 to December 1997, 386 patients with H. pylori infection received eradication treatments. The presence of H. pylori was assessed by rapid urease test, UBT, culture and histologic examination. Eradication of the bacteria was determined by the negative results in all of these four tests. At 4 weeks after completion of therapy, 312 cases (80.8%) were judged as being free of H. pylori. Mean observation period was up to 12 months, and 113 cases were followed up to more than 1 year, and 50 cases were followed up to more than 2 years. H. pylori had re-appeared in 3 cases after 3 months, 1 case after 6 months, 2 cases after 12 months, and 1 case 24 months after the treatment, respectively. For the purpose of more accurate diagnosis, the assessment of eradication of H. pylori should be performed at 1 year after the completion of therapy. Since all the recrudescence could be diagnosed with UBT earlier and be confirmed by the other tests later, UBT is recommended as a useful methods in the assessment of Helicobacter pylori eradication.

Detection methods of Helicobacter pylori: accuracy and costs.

A variety of methods exist for determining gastric colonization with Helicobacter pylori, which has been implicated in the development of peptic ulcer disease. The goal of this study was to evaluate four of the current methods available in a clinical surgical practice setting through a prospective evaluation of 40 consecutive patients undergoing upper diagnostic endoscopy. All patients underwent six antral gastric biopsies for use with the following detection methods: histologic demonstration of organisms (hematoxylin and eosin stain), direct detection of urease activity (Remel Selective Rapid Urea, Lenexa, KS), and culture of H. pylori. All patients also had measurement of serum immunoglobulin G for H. pylori by the enzyme-linked immunosorbent assay method (Corning Clinical Laboratories, St. Louis, MO). The infection status was established by a concordance of test results. The results show that H. pylori can be assessed equally well with histology, a rapid urease test, and serology, with all three tests having good sensitivity (92-100%) and specificity (85-96%). The culturing of the organism had poor sensitivity (42%). The benefits of the urease test are a much more rapid response time and a much lower cost as compared to histologic and serologic testing. In conclusion, the rapid urease test is the method of choice to detect H. pylori in those patients undergoing endoscopy in whom the identification of H. pylori will change their management.

Modification of Christensen urease test as an inexpensive tool for detection of Helicobacter pylori.

About half the world population is infected with Helicobacter pylori. Most live in developing countries where clinical studies face the constraints of high costs of imported rapid diagnostic tests. In this work, we describe and validate a simple local urease test (LUT) to determine the presence of the bacterium in gastric biopsies, and report the incidence of infection among symptomatic patients in Caracas, Venezuela. Statistical comparison of LUT and CLOtest (Delta West, Bentley, Australia) (N = 216 patients) showed that the probability of 95% agreement between the two test was 0.936. Overall incidence of infection determined by the LUT was 65% (N = 229), and it was higher in patients from public (72%; N = 153) than from private (50%; N = 76) hospitals (p = .001). Therefore, the incidence of infection differs in two socioeconomic groups that coexist in the same city. LUT may represent an affordable tool in clinical studies needed to identify social factors that increase the risk of infection by H. pylori.

The Pyloritek test and the CLO test: accuracy and incremental cost analysis.

OBJECTIVE: The aim of this study was to compare the Pyloritek test (a 1-h rapid urease test) to the widely used CLO test. METHODS: Seventy-one patients undergoing upper endoscopy were studied. All patients gave informed consent. A single antral biopsy specimen was obtained for the CLO test, and another was obtained for the Pyloritek test. Additional specimens were obtained for culture and processed in the event of a discordant result. The Pyloritek assay was read at 1 h by one observer. The CLO test was read at 24 h by an observer blinded to the results of the Pyloritek assay. RESULTS: There were 18 males and 53 females, and the mean age (+/- SEM) was 53 +/- 17 yr. Thirty-two patients had a positive result on the CLO test, and 39 had a negative test result. Of the 32 patients with a positive CLO test result at 24 h, 31 were positive by the Pyloritek test at 1 h. All 39 patients with negative CLO test results had negative Pyloritek test results as well. There was one discordant result, a negative Pyloritek test result and a positive CLO test result. Culture demonstrated growth of Helicobacter pylori. The kappa value, a measure of the reliability of the Pyloritek test compared with the CLO test, was 0.972 (SE, 0.0284; 95% confidence interval, 0.925-1). Marginal cost-effectiveness analysis favored the Pyloritek test. CONCLUSIONS: Results of the Pyloritek test at 1 h and the CLO test at 24 h are comparable in terms of detection of urease activity.

Clinical assessment of the bacterial load of Helicobacter pylori on gastric mucosa by a new multi-scaled rapid urease test.

The present study tests the efficacy of the multi-scaled urease test (MUT) in detecting Helicobacter pylori infection and determines whether the MUT can predict the bacterial density on histology. A total of 111 sets of gastric specimens were obtained from patients with dyspepsia but without recent bleeding. Two biopsies were taken as closely as possible in each set. One sample was used for the MUT (Hp fast; GI Supply, Camp Hill, PA, USA), while the other was used to determine the histological density of H. pylori by modified Giemsa stain (grade 0-5). The results of MUT were interpreted as negative if the colour was yellow or bright green (reaction score 0) and positive if the colour was green, light blue, or blue (reaction score 1, 2 and 3, respectively). The reaction scores of MUT were recorded sequentially at 15 and 30 min and 1, 4 and 24 h. On the basis of histological confirmation, MUT had a sensitivity of 89.6%, a specificity of 88.2%, a positive predictive value of 94.5% and a negative predictive value of 78.9%. Focusing on specimens with the presence of bacteria under histology, 77 specimens were divided into five subgroups by grades of density of H. pylori (HPD1-5). The reaction scores had become sequentially elevated from 30 min through to 24 h in each subgroup. For subgroups HPD4 and 5, the positive rates of MUT were 70.6 and 66.6%, respectively, as early as 30 min and progressed to 100% within 4 h. In contrast, the positive rate for the HPD1 subgroup was 16.6% at 4 h and increased to only 62.5% at 24 h. In subgroups HPD 2 and 3, the positive rates were less than 30% at 30 min, but became more than 66.6% at 4 h and were 100% at 24 h. The early (i.e. mean value of reaction scores before 4 h) and late (24 h) mean reaction scores disclosed two elevated trends as the density of H. pylori increased (early: 0.2, 0.7, 0.8, 1.5, 1.2; late: 1.4, 2.3, 2.6, 3.0, 3.0; P < 0.05). In conclusion, MUT is a reliable method for the diagnosis of H. pylori infection. It can also indirectly predict the density of H. pylori on histology.

Clinical utility and cost effectiveness of Helicobacter pylori testing for patients with duodenal and gastric ulcers.

OBJECTIVE: current consensus guidelines recommend that all patients demonstrating either a gastric or duodenal ulcer be tested for Helicobacter pylori, the most common cause of ulcers. We determined the clinical utility and cost effectiveness of H. pylori testing in patients with duodenal and gastric ulcers. METHODS: A retrospective evaluation and cost-effectiveness analysis of 565 consecutive patients with endoscopically demonstrated gastric or duodenal ulcers over a 4-yr period in a large, urban general hospital. The main outcome variables are the percentage of patients who had a gastric biopsy, the prevalence of H. pylori, and the cost effectiveness of testing (antral biopsy, CLO test, serum antibody, and urea breath test) for H. pylori. RESULTS: Significantly more patients with endoscopically documented duodenal ulcers had an antral biopsy performed in 1993 and 1994 when compared with patients from 1991 and 1992 (p < 0.00001). For patients with gastric ulcers, biopsies were performed at a similar rate throughout this study. Overall, patients with duodenal and gastric ulcers demonstrated H. pylori 75% and 69% of the time, respectively. The total charges for biopsy documentation and treatment of H. pylori in all duodenal ulcer patients in this cohort was estimated at $25,135. If a biopsy for H. pylori had been performed in all patients the actual charges would have been $77,443. Conversely, charges would have been only $8085 had all patients been empirically treated for H. pylori based on the high pretest probability of infection. CONCLUSIONS: Routine testing for H. pylori is very expensive, regardless of the diagnostic method used. Biopsy results do not provide clinically useful information in most patients with duodenal ulcers and may be misleading if falsely negative.

Assessment of four rapid urease test systems for detection of Helicobacter pylori in gastric biopsy specimens.

An in vitro experiment and an in vivo clinical trial were both performed in order to assess the efficacy of four biopsy urease test systems, including one commercial kit, Temmler CUT test (Temmler Pharma, Marburg, Germany), for the rapid detection of Helicobacter pylori. We first evaluated four rapid urease test systems by inoculating bacterial suspensions of different concentrations into urea-containing media and observing the color change at room temperature. We found that the CUT test was superior in vitro to the other three urease test systems. As was expected, the lower the concentration of the inoculum, the slower was the color change and the fewer were the positive results noted. The minimal concentration of H. pylori for a positive urease test at 24 h was 1000-10,000 colony-forming units/ml in 1 drop of bacterial suspension inoculated. We then evaluated four biopsy urease test systems for the rapid diagnosis of H. pylori infection in antral and fundal mucosa biopsy specimens of 37 patients undergoing upper gastrointestinal endoscopy. All four test systems were 100% specific when compared with culture. In 69 culture-positive biopsy specimens, the CUT test was not only more sensitive (72%) than the other three systems (42%, 51%, and 45%, respectively), but also gave the fastest reaction by detecting more culture-positive biopsy specimens after 3 h of incubation at room temperature. The differences were statistically significant.

Serum 13C-bicarbonate in the assessment of gastric Helicobacter pylori urease activity.

Noninvasive detection of Helicobacter pylori (HP) requires serum or salivary antibody testing or the CO2 breath test. Since gastric HP produces a potent urease, a meal rich in 13C-labeled urea should lead to a measurable quantity of isotopic CO2 in the serum. This study investigates the feasibility, sensitivity, specificity, and potential of the measurement of serum 13C-bicarbonate (13CHCO3) in determining the presence of gastric HP. Nineteen patients with upper gastrointestinal symptoms assessed by intensity-duration questionnaire underwent endoscopy and biopsies for histology, Giemsa stain, and urease activity testing by CLOtest. Patients also consumed a 13C-urea-rich meal (5 mg/kg body weight, 99% 13C, MSD Isotopes, Montreal Canada). Serum was collected every 30 min for 3 h for quantitative determination of 13C by mass spectrometry. Fractional elevation of 13C after the enriched meal was then correlated with endoscopy, histology, and CLOtest. Fourteen of the 19 patients studied had histologic evidence of gastritis; 11 of 19 had positive CLOtest and had HP by histology and Giemsa stain. All HP-positive patients had significant elevation of 13CHCO3, compared with HP-negative patients. The mean maximum absolute change from baseline was 15.3 delta 13CHCO3 (range, 6.7-29.9) and occurred from 15 to 90 min; 13CHCO3 in HP-negative patients was significantly less (p < 0.05) than HP-positive patients with a mean value of 2.3 delta 13CHCO3 (range, 0-5.3). We conclude that serum 13CHCO3 analysis accurately reflects HP gastritis. This novel method is noninvasive, less labor intensive, less time consuming, and may have a value as a diagnostic screening tool for humans or in the assessment of the results of therapy in patients with HP infection.

Field evaluation of a rapid, simple and inexpensive urease test for the detection of Helicobacter pylori.

A modified rapid urease test (MRU test) for the detection of Helicobacter pylori was evaluated under field conditions during an endoscopic survey in rural India and compared with a commercially available urease test (CLO test) and with histology. Of 195 consecutive subjects who underwent upper gastrointestinal endoscopy, 153 (78.5%) were positive for Helicobacter pylori when tested by the CLO test and/or histology. The sensitivity and specificity of the MRU test relative to this was 97.4 and 95.2%, respectively when the test was read over a 3 h period. The MRU test was positive in 77.4, 89.0, 93.8 and 96.6% of cases at 1, 5, 20 and 60 min, respectively, compared with 2.7, 14.4, 48.6 and 71.2% of cases for the CLO test at the same time. The accuracy of the MRU test was thus similar to that of other methods for the detection of Helicobacter pylori. Furthermore, it gave a positive diagnosis more rapidly than other tests, in most cases before the subject had left the endoscopy suite. The MRU test is extremely simple to prepare and read and costs less than 0.05 pounds per test compared with 2.26 pounds for a CLO test. It is suitable for use in clinical or epidemiological work and especially where cost factors are critical.

Use of an ammonia electrode for rapid quantification of Helicobacter pylori urease: its use in the endoscopy room and in the assessment of urease inhibition by bismuth subsalicylate.

The use of an ammonia electrode to quantify ammonia liberated by urease from Helicobacter pylori was assessed in an in vitro study. It was found to be highly sensitive (down to 0.7 ppm NH3) and highly reproducible (coefficient of variation 6.0%). Inhibition of urease by bismuth subsalicylate was evaluated as urease testing is often used to assess clearance of H. pylori in patients treated with bismuth. Concentrations of bismuth subsalicylate up to 5 mg/ml had no inhibitory effect but bismuth subsalicylate at 50 mg/ml resulted in 21% inhibition of the urease activity of an ultrasonicated H. pylori suspension. As a preliminary study, the ammonia electrode was assessed in the endoscopy room in comparison with conventional techniques for H. pylori diagnosis. Antral biopsies from 39 patients attending for routine diagnostic endoscopy were subjected to culture, histology, detection of urease activity with a commercially available slide test (CLO) and with the ammonia electrode to detect ammonia liberated from samples placed in urea solution. 21 patients were positive after 1 h with the ammonia electrode, compared to only 17 with the commercially available slide test. 20 were positive on histology and 19 by culture. All samples positive with the ammonia electrode were either positive by culture or by histology. The ammonia electrode offers a quick, sensitive, quantitative and cheap method for the detection and quantification of H. pylori.