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A C Frasch

Publications and source records attributed to A C Frasch.

At least 55 records · Page 3Linked to original sources
PubMed1994

Mice infected with Trypanosoma cruzi produce antibodies against the enzymatic domain of trans-sialidase that inhibit its activity.

trans-Sialidase (TS) is an enzymatic activity described only for trypanosomes that is involved in the invasion of host cells by Trypanosoma cruzi. The enzyme that is shed by the parasite is made of two domains, the C-terminal region containing immunodominant amino acid repeats that define the SAPA antigen and the N-terminal domain that contains the putative region for enzymatic activity. The SAPA antigen induces a strong humoral response detected shortly after infection, both in humans and in mice. This response is directed to the immunodominant domain but is irrelevant in terms of neutralization of TS activity. We now show that TS activity can be detected in sera from acutely infected mice. However, mice infected with a T. cruzi strain whose growth can be controlled by the host did not have detectable levels of TS activity in sera. In fact, sera from these mice were able to abolish TS activity. This inhibition was due to the presence of specific antibodies directed against the enzymatic domain of the protein since they also abolish the activity of a recombinant molecule lacking the immunodominant amino acid repeats. The neutralizing antibodies were present from day 30 after the infection, while antibodies to the immunodominant repeats were detected by day 8 postinoculation, suggesting that the in vivo role of these repeats is to defect the humoral response to the repeat domain until the infection is established.

Animals↗
PubMed1994

Direct detection of Vibrio cholerae in stool samples.

A direct method to detect Vibrio cholerae in stool samples was developed by using a PCR procedure that did not require a DNA purification step. Dilution (1/100) of stool samples prevented inhibition of the reaction by contaminants, and two consecutive PCRs, the second one with a nested primer, achieved the desired sensitivity. Comparison of the results obtained from stool swab samples processed by the two-step PCR and by an enzyme-linked immunosorbent assay using GM1 as the capture molecule showed that the former is more sensitive and gave positive results even when V. cholerae was not culturable or dead.

Argentina↗
PubMed1994

Immunoassay with recombinant Trypanosoma cruzi antigens potentially useful for screening donated blood and diagnosing Chagas disease.

A new enzyme immunoassay (EIA), the Dia Kit Bio-Chagas assay (Gador S.A.), is potentially useful for detecting antibodies to Trypanosoma cruzi in the diagnosis of infected individuals and the screening of blood in blood banks. The EIA is carried out on test strips of plastic backing covered with a nitrocellulose membrane to which a mixture of recombinant T. cruzi antigens 1, 2, shed acute-phase antigen, 13, and 30 has been applied as a horizontal line. A horizontal line of human IgG is included to monitor the test procedure. The test strips exhibited homogeneity in the adsorption of the mixture of recombinant antigens (CV = 6.0%) and the human IgG (CV = 8.6%). The EIA results obtained with sera positive by xenodiagnosis showed 100% agreement between both types of tests; tested against sera with positive and negative matched results of indirect hemagglutination, immunofluorescence, and ELISA, the EIA results agreed for 99.1% (347 of 350) and 99.6% (299 of 300) of the samples, respectively.

Adsorption↗
PubMed1993

The action of Trypanosoma cruzi trans-sialidase on glycolipids and glycoproteins.

Addition of sialic acid residues in the human pathogen Trypanosoma cruzi glycoconjugates is mediated by a trans-sialidase and not by a CMP-sialic acid:glycoconjugate sialyltransferase. Incubation of trans-sialidase with N-[galactose-14C]acetyllactosamine and O-linked oligosaccharides, N-linked glycopeptides (both obtained from fetuin) or sialyllactose showed that the last three compounds were donors of sialic acid residues to the first one. Moreover, N- and O-linked oligosaccharides in asialofetuin and asialomucin, respectively, served as acceptors of sialic acid units. Gangliosides GM3, GD1a and GT1b but not GM2, GM1a nor GD1b donated sialic acid units to N-acetyllactos amine when incubated with trans-sialidase. This showed that only sialic acid units bound to terminal galactosyl residues were transferred. GM1a was converted to GD1a, and GD1b to GT1b when incubated with the appropriate donor. The fact that asialo-GM1a was converted to a ganglioside migrating as GD1a on thin-layer chromatography suggested that sialic acid units may be transferred to internal galactosyl residues, although once linked to those residues they can not be further transferred to other glycoconjugates. Sialic acid residues linked alpha 2,3- but not alpha 2,6- or alpha 2,8- were transferred by the trans-sialidase. Methyl beta-galactoside but not methyl alpha-galactoside served as acceptor of sialic acid units, thus suggesting that terminal alpha-linked galactosyl units in T. cruzi and mammalian glycoproteins are not sialylated by the enzyme. As the trans-sialidase employed in these experiments has been shown to be located on the external surface of the parasite and to be shed to the medium, the relatively broad specificity shown by the enzyme with respect to protein- and lipid-linked oligosaccharides strongly suggests that infection by T. cruzi might alter the sialic acid distribution in glycoproteins and glycolipids of the mammalian host.

Animals↗
PubMed1993

Identification of toxigenic Vibrio cholerae from the Argentine outbreak by PCR for ctx A1 and ctx A2-B.

A polymerase chain reaction (PCR) to detect a region of the A1 cholera toxin gene was applied to the identification of 43 Vibrio cholerae strains isolated from the recent outbreak in Argentina. A good correlation was observed between the GM1-enzyme-linked immunosorbent assay (GM1-ELISA) to detect the B subunit of the enterotoxin and PCR. However, a V. cholerae non-01 strain that was negative by the ELISA test, was positive by the PCR assay for the A1 region. A second PCR test to detect the A2-B coding region was developed to solve this case. We propose that routine detection of toxigenic V. cholerae by PCR should include analysis of A2-B coding region or the whole cholera toxin operon.

Argentina↗
PubMed1993

The reactivity of sera from chagasic patients against different fragments of cruzipain, the major cysteine proteinase from Trypanosoma cruzi, suggests the presence of defined antigenic and catalytic domains.

Three fragments of cruzipain, expressed separately in bacterial vectors, were used to detect antibodies in sera from patients with chronic Chagas' disease. Most antibodies directed against the enzyme were found to react with the C-terminal extension, thus suggesting the presence of immunodominant B-cell epitopes within this protein domain. Immunoprecipitation with these antibodies did not impair enzyme activity. It is suggested that cruzipain consists of an enzymatic domain and a non-enzymatic, immunodominant domain, which corresponds to the C-terminal extension.

Animals↗
PubMed1992

Identification of the gene(s) coding for the trans-sialidase of Trypanosoma cruzi.

The gene(s) encoding the Trypanosoma cruzi shed-acute-phase-antigen (SAPA) has a 5' end encoding a region containing two totally and two partially conserved Ser-X-Asp-X-Gly-X-Thr-Trp motifs which are present in bacterial neuraminidases, and a 3' end encoding tandemly repeated units of 12 amino acids. It is now reported that 54-87% of the total neuraminidase activity present in the parasite could be immunoprecipitated with polyclonal or monoclonal antibodies against the repeated amino acid units of SAPA. These immunoprecipitates also had greater than 80% of the trans-sialidase activity of the parasite. SAPA used sialyllactose, fetuin and 4-methylumbelliferyl-sialic acid as substrate donors. In the presence of a suitable acceptor molecule (lactose) the sialic acid residues were transferred to the disaccharide, whereas in the absence of acceptors the residues were transferred to water. If relatively inefficient acceptors (maltose or cellobiose) were added to the incubation mixtures, the sialic acid units were transferred both to the disaccharides and to water. It is concluded that a major T. cruzi antigen has both the trans-sialidase and the neuraminidase activities of the parasite. Both activities are probably located on the N-terminus of SAPA since antibodies directed against the C-terminus, which contains the repeated amino acid units, do not affect the enzymatic activities.

Amino Acid Sequence↗
PubMed1992

The major cysteine proteinase (cruzipain) from Trypanosoma cruzi is encoded by multiple polymorphic tandemly organized genes located on different chromosomes.

We demonstrate that cruzipain, the major cysteine proteinase of Trypanosoma cruzi epimastigotes, is encoded by a large number of tandemly arranged genes. Restriction enzyme analysis of 20 clones containing complete repeat units of the gene, as well as sequencing of 2 of these clones, and comparison with previously published partial sequences, indicated that the sequence is conserved among the repeat units, although polymorphisms clearly exist. The repeat units contain an intergenic region of 528 bp and coding regions for pre- and pro-enzyme, a central domain and a C-terminal extension. The predicted amino acid sequences of these regions indicated a sequence identity of 30, 60, 70 and 36%, respectively, when the T. cruzi sequence was compared with the sequence of a similar cysteine proteinase from Trypanosoma brucei. Studies by pulsed field gel electrophoresis, complemented with restriction analysis, indicated that the clusters are located on 2-4 different chromosomes in several parasite isolates.

Amino Acid Sequence↗
PubMed1992

A superfamily of Trypanosoma cruzi surface antigens.

Several genes o f Trypanosoma cruzi encode surface antigens that include an amino acid motif that is conserved among bacterial neurominidases. Oscar Campetella, Daniel Sdnchez, Juan Jose Cazzulo and Alberto Carlos Frasch here suggest grouping these gene families in a superfamily.

Journal Article↗
PubMed1992

SAPA/trans-sialidase and cruzipain: two antigens from Trypanosoma cruzi contain immunodominant but enzymatically inactive domains.

Trypanosoma cruzi, the parasitic protozoan that causes the American trypanosomiasis, or Chagas disease, contains a number of antigenic molecules, some of which have tandems of amino acid repeats. One of these molecules, SAPA (shed acute phase antigen), contains a so-far unique trans-sialidase activity that is essential for penetration of the parasite into mammalian cells. The enzyme consists of two different domains, one presumably enzymatic, which contains four copies of an amino acid motif conserved in bacterial neuraminidases, and the other highly antigenic, consisting of the repeats. Another enzyme that seems to be involved in the host-parasite relationship, the cysteine proteinase cruzipain, is also made up in its mature form of a catalytic domain with high homology to cathepsin L and a COOH-terminal domain that is highly antigenic in vivo. These bifunctional molecules may have arisen by incorporation of a highly antigenic domain to an essential enzyme in order to attract the immune response, thus protecting the enzyme activity.

Amino Acid Sequence↗
PubMed1991

The expression of the major shed Trypanosoma cruzi antigen results from the developmentally-regulated transcription of a small gene family.

To better understand the mechanisms involved in the developmental expression of Trypanosoma cruzi antigens we examined the gene structure and transcription properties of the major shed trypomastigote antigen (SAPA). We report in this paper that SAPA is encoded by a small family of at lest 6 genes which differ mainly in the length of a repeat region made up of tandemly arranged 36-bp repeat units. SAPA genes are located distant from chromosomal telomeres as inferred from their insensitivity to Bal31 nuclease treatment. Furthermore, Northern blot and S1 protection analyses strongly support the fact that most (or all) SAPA genes are transcribed in the infective form of the parasite.

Animals↗
PubMed1991

Comparison of genes encoding Trypanosoma cruzi antigens.

Trypanosoma cruzi, the causative agent of Chagas disease, simultaneously expresses several different surface antigens. This contrasts sharply with blood-stream forms o f the African trypanosomes, which display only one variant surface glycoprotein at a time. Over the past few years, the genes coding for a number of T. cruzi proteins recognized by sera from patients have been cloned and at least partially sequenced. However, some of those discovered in more than one laboratory have been given different names. Here, Carlos Frasch, Juan Cazzulo, Lena Aslund and UIf Pettersson try to systematize the literature available on these antigens, including what is known about their localization and function.

Journal Article↗