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| 3. |
- Abu Al-Soud, Waleed, et al.
(författare)
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Assessment of PCR-DGGE for the identification of diverse Helicobacter species, and application to faecal samples from zoo animals to determine Helicobacter prevalence.
- 2003
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Ingår i: Journal of Medical Microbiology. - : Microbiology Society. - 0022-2615 .- 1473-5644. ; 52:9, s. 765-771
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Tidskriftsartikel (refereegranskat)abstract
- Helicobacter species are fastidious bacterial pathogens that are difficult to culture by standard methods. A PCR-denaturing gradient gel electrophoresis (PCR-DGGE) technique for detection and identification of different Helicobacter species was developed and evaluated. The method involves PCR detection of Helicobacter DNA by genus-specific primers that target 16S rDNA and subsequent differentiation of Helicobacter PCR products by use of DGGE. Strains are identified by comparing mobilities of unknown samples to those determined for reference strains; sequence analysis can also be performed on purified amplicons. Sixteen DGGE profiles were derived from 44 type and reference strains of 20 Helicobacter species, indicating the potential of this approach for resolving infection of a single host by multiple Helicobacter species. Some more highly related species were not differentiated whereas in highly heterogeneous species, sequence divergence was observed and more than one PCR-DGGE profile was obtained. Application of the PCR-DGGE method to DNA extracted from faeces of zoo animals revealed the presence of Helicobacter DNA in 13 of 16 samples; a correlation was seen between the mobility of PCR products in DGGE analysis and DNA sequencing. In combination, this indicated that zoo animals are colonized by a wide range of different Helicobacter species; seven animals appeared to be colonized by multiple Helicobacter species. By this approach, presumptive identifications were made of Helicobacter bilis and Helicobacter hepaticus in a Nile crocodile, Helicobacter cinaedi in a baboon and a red panda, and Helicobacter felis in a wolf and a Taiwan beauty snake. All of these PCR products (400 bp) showed 100 % sequence similarity to 16S rDNA sequences of the mentioned species. These results demonstrate the potential of PCR-DGGE-based analysis for identification of Helicobacter species in complex ecosystems, such as the gastrointestinal tract, and could contribute to a better understanding of the ecology of helicobacters and other pathogens with a complex aetiology.
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- Casswall, TH, et al.
(författare)
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Bovine anti-Helicobacter pylori antibodies for oral immunotherapy
- 2002
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Ingår i: Scandinavian Journal of Gastroenterology. - : Informa UK Limited. - 1502-7708 .- 0036-5521. ; 37:12, s. 1380-1385
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Tidskriftsartikel (refereegranskat)abstract
- Background: Passive immunization with orally administered antibodies against specific pathogens has previously been successfully used therapeutically in both animal and human studies. We employed a similar strategy for experimental treatment of mice infected with the gastric pathogen Helicobacter pylori. Methods: An anti-H. pylori bovine colostral hyperimmune immunoglobulin preparation (BIC) was generated and its efficacy was tested in different in vitro experiments, such as binding to the Lewis(b) blood group antigen, inhibition of adherence of H. pylori to human gastric mucosa tissue sections in situ and in a haemagglutination assay. The BIC preparation was also given in the drinking water to H. pylori-infected mice. Results: An inhibition of 95% of the binding of H. pylori to Lewis(b) glycoconjugate was observed in vitro. Furthermore, a blocking activity of almost 90% was observed when the BIC was preincubated with H. pylori bacteria. Finally, the BIC preparation inhibited the haemagglutination of H. pylori and human red blood cells. Seven of 40 (17.5%) mice remained infected in the treatment group as compared with 25 of 45 (55.5%) in the control group. Hence, the cure rate was 66%, P = < 0.001. The mean number of colonies in the antibody-treated mice where eradication was not successful was also reduced (P < 0.05). In trials using FVB/N transgenic Lewis(b) expressing mice, a cure rate of 50%-66% was observed. Conclusion: Bovine colostral antibodies against H. pylori can be generated in high titres, inhibit binding in vitro and can eradicate or reduce the number of bacteria in infected mice.
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- Drangert, Jan-Olof, et al.
(författare)
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Why Did They Become Pipe-Bound Cities? : Early Water and Sewerage Alternatives in Swedish Cities
- 2002
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Ingår i: Public Works Management & Policy. - : Sage Publications. - 1087-724X .- 1552-7549. ; 6:3, s. 172-172-185
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Tidskriftsartikel (refereegranskat)abstract
- For decades and even centuries, Swedish towns, prompted by the need for clean water, haddiscussedpotential water systems. Some towns haddevelopedplans, but it was first in the secondhalf of the 19th century that the gradual conversion was made to pipe-boundsystems for water andsanitation. Not until 1863, after towns hadgainedthe authority to collect taxes, borrow money, lay pipes through private compounds, and monitor the urban environment, did infrastructure development become the task of local government. Cholera epidemics and fire protection (and thus lower insurance fees) were among the factors motivating the town councils andproperty owners. Especially from 1875 on, the hygienic value of water was also emphasizedin the effort to enforce public health legislation. Pipes for the systems were importedanddesigns emulated.
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- Jideus, Lena, et al.
(författare)
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Thoracic epidural anesthesia does not influence the occurrence of postoperative sustained atrial fibrillation
- 2001
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Ingår i: Annals of Thoracic Surgery. - 0003-4975 .- 1552-6259. ; 72:1, s. 65-71
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Tidskriftsartikel (refereegranskat)abstract
- Background. To evaluate whether thoracic epidural anesthesia (TEA) can reduce the incidence of atrial fibrillation (AF) after coronary artery bypass grafting (CABG).Methods. Forty-one patients undergoing CABG were treated with TEA intraoperatively and postoperatively. Another 80 patients served as the control group. The sympathetic and parasympathetic activities were evaluated by analysis of neuropeptides, catecholamines and heart rate variability (HRV), preoperatively and postoperatively.Results. Postoperative AF occurred in 31.7% of the TEA-treated patients and in 36.3% of the untreated patients (p = 0.77). TEA significantly suppressed sympathetic activity, as indicated by a less pronounced increase of norepinephrine and epinephrine (p = 0.03, p = 0.02) and a significant decrease of neuropeptide Y (p = 0.01) postoperatively in TEA-treated patients compared to untreated patients. The HRV variable expressing sympathetic activity was significantly lower and the postoperative increase in heart rate was significantly less in the TEA group than in the control group after surgery (p = 0.01, p < 0.001). Among patients developing AF, the maximal number of supraventricular premature beats per minute increased significantly in untreated patients postoperatively but remained unchanged in TEA-treated patients (p = 0.004 versus p = 0.86).Conclusions. TEA has no effect on the incidence of postoperative sustained AF, despite a significant reduction in sympathetic activity.
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- Nelson, Marie C, 1944-, et al.
(författare)
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Why did they become pipe-bound cities?
- 2002
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Ingår i: Public Works Management & Policy. - 1087-724X .- 1552-7549. ; 6:3, s. 172-185
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Tidskriftsartikel (refereegranskat)
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| 8. |
- Nilsson, Anna, et al.
(författare)
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Partitioning of peptide-tagged proteins in aqueous two-phase systems using hydrophobically modified micelle-forming thermoseparating polymer
- 2002
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Ingår i: Biochimica et Biophysica Acta - Proteins and Proteomics. - 1570-9639. ; 1601:2, s. 138-148
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Tidskriftsartikel (refereegranskat)abstract
- Genetic engineering has been used to construct hydrophobically modified fusion proteins of cutinase from Fusarium solani pisi and tryptophan-containing peptides. The aim was to enhance the partitioning of the tagged protein in a novel aqueous two-phase system formed by only one water-soluble polymer. The system was based on a hydrophobically modified random copolymer of ethylene oxide (EO) and propylene oxide (PO) units, HM-EOPO, with myristyl groups (C14H29) at both ends. The HM-EOPO polymer is strongly self-associating and has a lower critical solution temperature (cloud point) at 12oC in water. At temperatures above the cloud point a two-phase system is formed with a water top phase and a polymer-enriched bottom phase. By adding a few percent of hydroxypropyl starch polymer, Reppal PES 200, to the system, it is possible to change the densities of the phases so the HM-EOPO-enriched phase becomes the top phase and Reppal-enriched phase is the bottom phase. Tryptophan-based peptides strongly preferred the HM-EOPO rich phase. The partitioning was increased with increasing length of the peptides. Full effect of the tag as calculated from peptide partitioning data was not found in the protein partitioning. When a short spacer was introduced between the protein and the tag the partitioning was increased, indicating a better exposure to the hydrophobic core of the polymer micelle. By adding a hydrophilic spacer between the protein and trp-tag, it was possible to increase the partitioning of cutinase 10 times compared to wild-type cutinase partitioning. By lowering the pH of the system and addition of NaCl, the partitioning of tagged protein was further increased towards the HM-EOPO phase. After isolating the HM-EOPO phase, the temperature was increased and the protein was back-extracted from the HM-EOPO phase to a fresh water phase.
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| 9. |
- Nilsson, Anna, et al.
(författare)
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Tryptophan-tagged cutinase studied by steady state fluorescence for understanding of tag interactions in aqueous two-phase systems
- 2003
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Ingår i: Biochimica et Biophysica Acta - Proteins and Proteomics. - 1570-9639. ; 1646:1-2, s. 57-66
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Tidskriftsartikel (refereegranskat)abstract
- Genetic engineering has been used to construct fusion proteins of Fusarium solani pisi cutinase and tryptophan-based tags, expressed in Saccharomyces cerevisiae, to increase the partitioning in aqueous two-phase systems. The separation systems were composed of thermoseparating polymers (random copolymers of ethylene oxide and propylene oxide, EOPO) and detergents (C12EOn). In this study, the fluorescence behaviour of the peptide-tagged protein, free peptide tag and tryptophan was investigated. The tryptophan-tagged proteins, cutinase-(WP)4 and cutinase-TGGSGG-(WP)4, showed emission spectra similar to the free peptides and tryptophan, indicating solvent exposure of the tag. The influence of polymers and detergents on the fluorescence of tagged proteins was examined. When peptides and tagged proteins were exposed to polymer, a slight blue shift of the emission maximum was observed. Larger blue shifts of the emission maximum were observed when C12EOn detergents were utilised. The results correlate with aqueous two-phase partitioning where addition of C12EOn detergents results in more extreme partitioning compared to systems containing only polymers. Dynamic light scattering (DLS) measurements of the EOPO copolymers were carried out, showing that the polymers did not aggregate at concentrations used in aqueous two-phase systems. Quenching of fluorescence with iodide for both proteins and peptide tags was studied. Plots according to the Stern-Volmer equation resulted in a linear fit, indicating exposed tryptophan residues for both free peptides and fusion proteins. The quenching constants were similar for both tagged protein and free peptide tag. The fluorescence results indicated that the tryptophan residues in the tag were exposed to the solvent and could interact with detergents and polymers in the two-phase systems.
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| 10. |
- Nilsson, Hans-Olof, et al.
(författare)
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Effect of Cold Starvation, Acid Stress, and Nutrients on Metabolic Activity of Helicobacter pylori.
- 2002
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Ingår i: Applied and Environmental Microbiology. - 0099-2240. ; 68:1, s. 11-19
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Tidskriftsartikel (refereegranskat)abstract
- Helicobacter pylori can transform, in vivo as well as in vitro, from dividing spiral-shaped forms into nonculturable coccoids, with intermediate forms called U forms. The importance of nonculturable coccoid forms of H. pylori in disease transmission and antibiotic treatment failures is unclear. Metabolic activities of actively growing as well as nonculturable H. pylori were investigated by comparing the concentrations of cellular ATP and total RNA, gene expression, presence of cytoplasmic polyphosphate granules and iron inclusions, and cellular morphology during extended broth culture and nutritional cold starvation. In addition, the effect of exposing broth-cultured or cold-starved cells to a nutrient-rich or acidic environment on the metabolic activities was investigated. ATP was detectable up to 14 days and for at least 25 days after transformation from the spiral form to the coccoid form or U form in broth-cultured and cold-starved cells, respectively. mRNAs of VacA, a 26-kDa protein, and urease A were detected by using reverse transcription-PCR in cells cultured for 2 months in broth or cold starved for at least 28 months. The ATP concentration was not affected during exposure to fresh or acidified broth, while 4- to 12-h exposures of nonculturable cells to lysed human erythrocytes increased cellular ATP 12- to 150-fold. Incubation of nonculturable cold-starved cells with an erythrocyte lysate increased total RNA expression and ureA mRNA transcription as measured by quantitative real-time reverse transcription-PCR. Furthermore, the number of structurally intact starved coccoids containing polyphosphate granules increased almost fourfold (P = 0.0022) under the same conditions. In conclusion, a specific environmental stimulus can induce ATP, polyphosphate, and RNA metabolism in nonculturable H. pylori, indicating viability of such morphological forms.
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