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Biomedical subjects

G Wiedermann

Publications and source records attributed to G Wiedermann.

At least 37 records · Page 2Linked to original sources
PubMed1997

Molecular and biochemical characterization of phosphoglucomutases from Entamoeba histolytica and Entamoeba dispar.

Entamoeba histolytica and Entamoeba dispar have only recently been defined as two separate species. E. histolytica, the pathogenic species, is the microorganism causing invasive intestinal amoebiasis and/or liver abscess, while the morphologically similar E. dispar is nonpathogenic and noninvasive. The gold standard for the distinction of the two species has been the isoenzyme electrophoresis of phosphoglucomutases (EC 5.4.2.2) and hexokinases (EC 2.7.1.1), but there had also been a controversy about the possibility of a conversion of isoenzyme patterns. In this study, we cloned the phosphoglucomutase (PGM) cDNAs from the pathogenic and the nonpathogenic species. The deduced amino acid sequences were only 2.4% different. The cDNAs were expressed in Escherichia coli under the control of a T7 RNA polymerase promoter. The recombinant polypeptides displayed strong phosphoglucomutase activity, each of the recombinant enzymes comigrated with its natural counterpart from E. histolytica and E. dispar in the starch gel electrophoresis. Our results give a biochemical interpretation of the PGM isoenzyme pattern and support the clear distinction between the two species.

Amino Acid Sequence↗
PubMed1997

Protection against invasive amebiasis by a single monoclonal antibody directed against a lipophosphoglycan antigen localized on the surface of Entamoeba histolytica.

A panel of monoclonal antibodies was raised from mice immunized with a membrane preparation from Entamoeba histolytica, the pathogenic species causing invasive amebiasis in humans. Antibody EH5 gave a polydisperse band in immunoblots from membrane preparations from different E. histolytica strains, and a much weaker signal from two strains of the nonpathogenic species Entamoeba dispar. Although the exact chemical structure of the EH5 antigen is not yet known, the ability of the antigen to be metabolically radiolabeled with [32P]phosphate or [3H]glucose, its sensitivity to digestion by mild acid and phosphatidylinositol-specific phospholipase C, and its specific extraction from E. histolytica trophozoites by a method used to prepare lipophosphoglycans from Leishmania showed that it could be classified as an amebal lipophosphoglycan. Confocal immunofluorescence and immunogold labeling of trophozoites localized the antigen on the outer face of the plasma membrane and on the inner face of internal vesicle membranes. Antibody EH5 strongly agglutinated amebas in a similar way to concanavalin A (Con A), and Con A bound to immunoaffinity-purified EH5 antigen. Therefore, surface lipophosphoglycans may play an important role in the preferential agglutination of pathogenic amebas by Con A. The protective ability of antibody EH5 was tested in a passive immunization experiment in a severe combined immunodeficient (SCID) mouse model. Intrahepatic challenge of animals after administration of an isotype-matched control antibody or without treatment led to the development of a liver abscess in all cases, whereas 11 out of 12 animals immunized with the EH5 antibody developed no liver abscess. Our results demonstrate the importance and, for the first time, the protective capacity of glycan antigens on the surface of the amebas.

Animals↗
PubMed1997

Molecular biology of the hexokinase isoenzyme pattern that distinguishes pathogenic Entamoeba histolytica from nonpathogenic Entamoeba dispar.

The electrophoretic patterns of hexokinase and phosphoglucomutase have been widely used to distinguish Entamoeba histolytica from Entamoeba dispar isolates. Although E. histolytica and E. dispar, previously called pathogenic and nonpathogenic Entamoeba histolytica, differ clearly in sequences of many homologous genes, a conversion between the two has been reported by several laboratories, in each case showing the conversion of hexokinase (ATP, D-hexose 6-phosphotransferase, EC 2.7.1.1) isoenzyme patterns. An apparent mobility shift of this enzyme may either be due to posttranslational modification or processing, or to the appearance of a new isoform encoded by a second gene. In this study we observed that the four observed bands in the isoenzyme patterns of pathogenic and nonpathogenic forms of Entamoeba were correlated with four different cDNAs, and that the four recombinant hexokinases produced in Escherichia coli comigrated with their natural counterparts. Polymerase chain reaction (PCR) experiments did not reveal hidden genes which might be responsible for conversion phenomena. These results strongly support the redefinition of pathogenic and nonpathogenic Entamoeba histolytica as two closely related species Entamoeba histolytica and Entamoeba dispar.

Amino Acid Sequence↗
PubMed1997

Immunogenicity and protectivity of a new liposomal hepatitis A vaccine.

In this study we investigated a new liposomal hepatitis A vaccine (Epaxal) developed by the Swiss Serum and Vaccine Institute clinically and immunologically using a one dose priming schedule and a booster injection after 1 year. This vaccine contains formalin inactivated hepatitis A virus particles attached to phospholipid vesicles together with influenza virus haemagglutinin. Two doses of the vaccine were administered at months 0 and 12 in 117 volunteers. Blood samples were drawn at days-7, 14 and 28 and after 6, 12 and 13 months, local and systemic reactions were monitored by means of questionnaires. Immunogenicity was evaluated as usual by the determination of anti-HAV from the collected sera using the ELISA technique. In order to evaluate the protective efficacy of the vaccine induced antibodies a sample of 25 sera mainly from vaccinees showing low ELISA titres was additionally analysed by means of a virus NT. The vaccine was excellently tolerable and highly immunogenic. Seroconversion evaluated by ELISA was 97 and 99%, respectively, 14 and 28 days after the first dose and 100% after the second dose. NT titres were well correlated with ELISA titres and showed similar seroconversion rates even in the early phase of immunization. The results of this study show that with two doses of the liposomal hepatitis A vaccine administered at months 0 and 12 early protection within 14 days and long lasting immunity can be achieved.

Adult↗
PubMed1997

Safety and immunogenicity of live oral cholera and typhoid vaccines administered alone or in combination with antimalarial drugs, oral polio vaccine, or yellow fever vaccine.

The effects of concomitant administration of antimalarial drugs, oral polio vaccine, or yellow fever vaccine on the immune response elicited by the Vibrio cholerae CVD103-HgR and Salmonella typhi Ty21a live oral vaccines were investigated. Healthy adults were immunized with CVD103-HgR alone or combined with Ty21a. Subjects were randomized to simultaneously receive mefloquine, chloroquine or proguanil, or oral polio or yellow fever vaccine. The vibriocidal antibody seroconversion rate was significantly reduced (P = .008) only in the group that received chloroquine with the CVD103-HgR. The geometric mean vibriocidal antibody titer was significantly decreased in the groups that received chloroquine (P = .001) or mefloquine (P = .02) compared with titers in groups that received CVD103-HgR alone. However, similar immunosuppressive effects were not observed in the groups immunized with Ty21a and CVD103-HgR. Only the concomitant administration of proguanil effected a significant (P = .013) decline in the anti-S. typhi lipopolysaccharide antibody response. These results indicate that chloroquine and proguanil should not be simultaneously administered with the CVD103-HgR and Ty21a vaccine strains, respectively.

Administration, Oral↗
PubMed1996

The sequence and organization of the core histone H3 and H4 genes in the early branching amitochondriate protist Trichomonas vaginalis.

Among the unicellular protists, several of which are parasitic, some of the most divergent eukaryotic species are found. The evolutionary distances between protists are so large that even slowly evolving proteins like histones are strongly divergent. In this study we isolated cDNA and genomic histone H3 and H4 clones from Trichomonas vaginalis. Two histone H3 and three histone H4 genes were detected on three genomic clones with one complete H3 and two complete H4 sequences. H3 and H4 genes were divergently transcribed with very short intergenic regions of only 194 bp, which contained T. vaginalis-specific as well as histone-specific putative promoter elements. Southern blot analysis showed that there may be several more histone gene pairs. The two complete histone H4 genes were different on the nucleotide level but encoded the same amino acid sequence. Comparison of the amino acid sequences of the T. vaginalis H3 and H4 histones with sequences from animals, fungi, and plants as well as other protists revealed a significant divergence not only from the sequences in multicellular organisms but especially from the sequences in other protists like Entamoeba histolytica, Trypanosoma cruzi, and Leishmania infantum.

Amino Acid Sequence↗
PubMed1996

Randomized, double-blind placebo-controlled trial to evaluate the safety and immunogenicity of combined Salmonella typhi Ty21a and Vibrio cholerae CVD 103-HgR live oral vaccines.

Healthy adults (n=330) were randomized to receive either a bivalent vaccine composed of Vibrio cholerae CVD 103-HgR and Salmonella typhi Ty21a or a placebo. The combined vaccine was well tolerated. Approximately 80% of vaccines manifested a significant rise in anti-S. typhi immunoglobulin G or immunoglobulin A lipopolysaccharide antibody levels. Significant (fourfold or greater) rises in anti-Inaba or anti-Ogawa vibriocidal antibody titer were achieved by 94 and 80% of vaccine recipients, respectively. Elevated baseline vibriocidal antibody titers showed a modest suppressive effect on the rate of seroconversion.

Administration, Oral↗
PubMed1996

Pyruvate: ferredoxin oxidoreductase from Entamoeba histolytica recognized by a monoclonal antibody.

A mouse monoclonal antibody, Eh208C2-2 MAb, raised against whole cell antigens of Entamoeba histolytica trophozoites of the pathogenic strain HM-1: IMSS and polyclonal antisera (PAb) against membrane antigens of E. histolytica trophozoites of strain HTH-56: MUTM were screened against a cDNA library of the pathogenic strain, SFL3. The monoconal antibody detected many phage plaques expressing an E. histolytica protein. The DNA sequence encoding the protein was approximately 55% identical, over 1,100bp, to Trichomonas vaginalis pyruvate: ferredoxin oxidoreductase (PFOR) and pyruvate: flavodoxin oxidoreductase from Klebsiella pneumoniae, Anabaena variabilis and Enterobacter agglomerans. Two of seven clones detected by mouse polyclonal antisera also encoded this protein. Two others encoded Entamoeba Hsp70, another encoded Entamoeba alkyl-hydroperoxide reductase and the remaining two were unidentified sequences. Entamoeba PFOR is an abundant, antigenic protein which may be a useful target for the development of protective host immune responses against invasive amebiasis.

Amino Acid Sequence↗
PubMed1995

The basic isoform of profilin in pathogenic Entamoeba histolytica. cDNA cloning, heterologous expression, and actin-binding properties.

In the human parasite Entamoeba histolytica, components of the cytoskeleton are involved in the pathogenicity by their contribution to immune evasion by antibody capping and shedding. In this study, we focus on profilin as a central regulatory component of the cytoskeleton. Profilin was isolated from trophozoites of the pathogenic E. histolytica strain SFL-3, and partial amino acid sequences were used to devise a probe for isolating a profilin cDNA. The deduced complete primary structure was divergent: plant profilins with amino acid sequence identities in the range 33-38% were more closely related than the mammalian profilins with sequence identities 21-28%. The cDNA was expressed as a nonfusion protein in Escherichia coli. Isoelectric focussing of the natural profilin isolated from E. histolytica showed two isoforms with different isoelectric points; the recombinant profilin migrated with the basic isoform. In a blot overlay experiment, purified 125I-labeled recombinant profilin bound not only to plant actin, but also to mammalian actin, demonstrating that cytoskeletal components from distantly related organisms with divergent primary structures can be compatible.

Actins↗
PubMed1995

Molecular analysis of two hexokinase isoenzymes from Entamoeba histolytica.

The zymodemes, electrophoretic patterns of hexokinase, phosphoglucomutase and glucose phosphate isomerase isoenzymes, have been widely used to determine the pathogenicity of Entamoeba histolytica isolates. Although pathogenic and nonpathogenic forms of E. histolytica differ clearly in sequences of many homologous genes, a conversion between pathogenic and nonpathogenic zymodemes has been reported by several laboratories. To approach the question what might be the basis for the observed conversion, we examined the molecular biology of the hexokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1) isoenzymes in pathogenic E. histolytica. We isolated two different cDNAs pHXK1 and pHXK2 coding for polypeptides with significant sequence similarity to hexokinases and deduced molecular masses of 49.8 kDa and 49.4 kDa. The two hexokinase sequences differed by 11% on the amino acid and by 8% on the nucleotide level. Expression of the cDNAs in Escherichia coli as nonfusion proteins gave two polypeptides with hexokinase activity. The recombinant Hxk1 and Hxk2 polypeptides comigrated with the more basic and more acidic isoforms of pathogenic amoebae in starch gel electrophoresis, as well as in low and high resolution isoelectric focussing gels. This identified the observed hexokinase isoenzymes of pathogenic E. histolytica as the products of two genes, hxk1 and hxk2.

Amino Acid Sequence↗
PubMed1995

Safety and immunogenicity of a live oral bivalent typhoid fever (Salmonella typhi Ty21a)-cholera (Vibrio cholerae CVD 103-HgR) vaccine in healthy adults.

The safety and immunogenicity of the live oral attenuated vaccine strains vibrio cholerae CVD 103-HgR and Salmonella typhi Ty21a were evaluated alone or in a combined bivalent formulation in four groups composed of 185 healthy European adults. All presentations were well tolerated. The serum anti-S. typhi lipopolysaccharide immunoglobulin G and immunoglobulin A antibody responses were comparable for all groups (66 to 72% seroconversion). The serum vibriocidal antibody seroconversion rate ranged from 78 to 92.5% (P > 0.05) among the groups. However, the peak and geometric mean vibriocidal antibody titers were significantly higher (P < 0.005) in the groups which received the bivalent formulation along with two doses of Ty21a than in the group which received CVD 103-HgR followed by two doses of killed Escherichia coli K-12 placebo. The ingestion of a placebo shortly after CVD 103-HgR may have suppressed the magnitude of the immune response. These findings demonstrate the feasibility of producing multivalent live oral attenuated vaccines.

Administration, Oral↗
PubMed1994

Clinical and immunological investigation of a new combined hepatitis A and hepatitis B vaccine.

As with hepatitis B vaccines, the recently developed hepatitis A vaccine is suitable not only for individual protection, but also for public health control measures. For introduction into routine immunisation programmes, however, hepatitis A vaccine should preferably be combined with other already established vaccines. In particular, a combination of hepatitis A and hepatitis B vaccines would be appropriate. We investigated a new combined hepatitis A/hepatitis B vaccine comparing its tolerability and immunogenicity with that obtained after separate or mixed simultaneous administration of the two components. Three groups of healthy volunteers, each of approximately 50 persons, were included. All were negative for hepatitis A and hepatitis B markers and had normal liver enzyme values. They received hepatitis A (720 ELISA units) and hepatitis B (20 micrograms) vaccines in the deltoid muscle, combined, mixed or separately, according to a 0, 1, 6-month schedule. Blood samples for determination of antibodies to hepatitis A virus (anti-HAV) and hepatitis B virus (anti-HBs) and of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were drawn at months 0, 1, 2, 6, and 7. Local and systemic reactions were monitored by means of questionnaires. The results of our study demonstrate that the combined hepatitis A and B vaccine is well tolerated and highly immunogenic. The seropositivity and seroprotection rates were 100% for both antigens in all groups. Surprisingly, anti-HAV and anti-HBs antibody titres after the combined and mixed vaccines were significantly higher compared with the respective monovalent vaccines injected separately.

Adult↗