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G F Mitchell

Publications and source records attributed to G F Mitchell.

At least 91 records · Page 5Linked to original sources
PubMed1988

Molecular and serological characteristics of the glutathione S-transferases of Schistosoma japonicum and Schistosoma mansoni.

When aqueous extracts of Schistosoma japonicum and S. mansoni adult worms are passed over columns of glutathione-conjugated agarose, two molecular species of Mr 26,000 and Mr 28,000 are detected in eluates as analysed by SDS-PAGE, these eluates having glutathione S-transferase (GST) activity. The molecules, termed Sj26 and Sj28 from S. japonicum and Sm26 and Sm28 from S. mansoni, can be immunogenic in rabbits or mice and appear not to be linked together as subunits of GST heterodimers. The elution profile of SjGST (Sj26+Sj28) from glutathione columns resembles that of SmGST (Sm26+Sm28) and, by peptide mapping, radioiodinated Sj26 and Sm26 are related as are the two Mr 28,000 molecules. Similarities between radioiodinated Sj28 and Sm28 are also obvious on two-dimensional gel electrophoresis with some differences being observed between Sj26 and Sm26. The Mr 28,000 molecules are more prominent than the Mr 26,000 molecules and, although Sj28 and Sm28 is a poor immunogen in mice, immunological cross-reactivity between Sj28 and Sm28 is generally more readily detected than that between Sj26 and Sm26. Whether experimental vaccination against schistosomiasis japonica and schistosomiasis mansoni reported with cloned GSTs can be improved by incorporation of both Mr 28,000 and Mr 26,000 species into the vaccine remains to be determined. On this point, the present data suggest that vaccination of mice with Sj26 plus Sm28 should be a useful means of increasing antibody responses to the GSTs of S. japonicum.

Animals↗
PubMed1988

Schistosoma mansoni antigens differentially recognized by resistant WEHI 129/J mice.

Mice of the strain WEHI 129/J are genetically resistant to chronic Schistosoma mansoni infection. Resistance is expressed in at least 50% of mice, with the remaining mice showing normal susceptibility to infection. The serum antibody specificities in the resistant proportion of WEHI 129/J were analyzed at various times after exposure to cercariae by using both Western blotting and immunoprecipitation. Comparisons with the susceptible proportion of WEHI 129/J and other permissive mouse strains revealed four antigens that were differentially recognized by resistant mice at various times of infection: Sm25, an Mr 25,000 integral membrane protein of adult worms that was better recognized by resistant mice 40 to 50 days after exposure; Sm67, an Mr 67,000 water-soluble antigen of adult worms that was better recognized by resistant mice at days 30 to 40; Sm120, an Mr 120,000 antigen expressed by cercariae and adult worms that was differentially recognized, although inconsistently, at days 20 to 40 postexposure; and Sm26, an Mr 26,000 glutathione S-transferase that was uniquely recognized by resistant mice at day 20 in two of three experiments. Analysis of antibody specificities in (BALB/c x WEHI 129/J)F1 x WEHI 129/J backcross mice indicated that high responsiveness to Sm25 at days 40 to 50 correlated with resistance. The candidacy of these four molecules as vaccines for schistosomiasis mansoni is discussed.

Animals↗
PubMed1988

Schistosoma mansoni and S. japonicum worm numbers in 129/J mice of two types and dominance of susceptibility in F1 hybrids.

In a study on the genetics of resistance to schistosomiasis in WEHI 129/J mice, susceptibility to either Schistosoma mansoni or Schistosoma japonicum was shown to be unequivocally dominant in F1 hybrid crosses between genetically resistant WEHI 129/J and susceptible BALB/c mice. The operation of only 1 or 2 genes in the expression of resistance to S. mansoni was suggested by backcross analysis. Thus, approximately 25% of (BALB/c x WEHI 129/J) F1 x WEHI 129/J mice were resistant to S. mansoni infection, whereas resistance was manifest in approximately 50% of WEHI 129/J mice. The data are consistent with resistance being controlled by 1 recessive gene having 50% penetrance. We also report that 129/J mice obtained directly from the Jackson Laboratories (Bar Harbor, Maine) (designated JAX 129/J), differ from locally bred WEHI 129/J in being entirely susceptible to S. mansoni infection. However, both WEHI 129/J and JAX 129/J are relatively resistant to S. japonicum infection.

Animals↗
PubMed1988

Immunoparasitology: its contribution to development of new parasite vaccines.

Immunoparasitology--the study of the immunology of host-parasite relationships--can post some notable research successes over the past decade. Progress towards prophylactic molecularly-defined vaccines against human parasitic diseases such as falciparum malaria, schistosomiasis mansoni and cutaneous leishmaniasis, as well as economically-important veterinary parasites, has been good. However, new vaccines are not coming as easily as might be hoped mainly because of several deficiencies in knowledge on the immunology of host-parasite relationships and the unknown relevance of well-characterized model systems to real-life parasitic diseases. In some models, the immunology of resistance and the immunology of disease are understood in broad outline. The availability of isolated antigens and their epitopes has improved quantitation of host immune responses to various life cycle stages of parasites and enabled vaccination efficacy or diagnostic potential to be assessed. One of several major challenges facing the immunoparasitologist interested in vaccine development is overcoming genetically-based unresponsiveness to "oligoepitope", defined-antigen vaccines particularly at the level of helper TH and cytotoxic (Tc) T cells.

Animals↗
PubMed1988

Analysis of variables associated with promotion of resistance and its abrogation in T cell-reconstituted nude mice infected with Leishmania major.

Upon intradermal challenge with the protozoan parasite Leishmania major, some mouse strains develop chronic cutaneous lesions, whereas other mouse strains show a resolving pattern of disease. The importance of T cell-dependent immunity in resistance to cutaneous leishmaniasis is substantiated by the susceptibility to infection of athymic nude mice of both resistant and susceptible strains. Small numbers of T lymphocytes from uninfected euthymic mice promote resistance in nude mice but T cells from chronically infected mice can impair this protective effect. In the present study we used an adoptive transfer system in which nude mice were reconstituted with T cells from normal or chronically infected mice in order to further investigate protection against disease or disease promotion. The results supported the following conclusions: (a) the host-protective activity of T cells from uninfected mice is highly effective even in long-term chronically infected nude mice, (b) T cell-mediated exacerbation of cutaneous disease does not involve enhancement of lesion development and is thus unlikely to be based on an accelerated proliferation of parasites in the lesion, (c) disease-promoting cells are not only found in genetically susceptible mice but can also be induced in genetically resistant mice, and (d) lymphoid organs of genetically susceptible mice chronically infected with L. major contain resistance-promoting cells in addition to disease-promoting cells. The data, together with those of others, continue to support the notion that recruitment with expansion and/or activation of different T cell subsets underlies genetically based resistance and susceptibility of mice to L. major.

Animals↗
PubMed1987

Injection versus infection: the cellular immunology of parasitism.

Parasites are immunogenic. But the variety of immune responses they can elicit is matched by their range of mechanisms to evade, subvert or distract these responses. The parasites' aim is survival and reproduction, ours is to restrict or eliminate them, and amplification of protective immune responses has become a key approach to this. But while vaccination has achieved many successes against the 'simpler' organisms such as viruses, the more complex protozoa and helminth parasites have proven much less tractable. In this article, Graham Mitchell discusses the differences between immune responses operated by whole parasites and those induced by prepared fractions.

Journal Article↗
PubMed1987

Heterologous protection in murine cutaneous leishmaniasis.

Mice immunized with a glycolipid antigen (GL) of Leishmania major plus adjuvant are relatively resistant to subsequent infection with this protozoan parasite. The GL is affinity purified on the monoclonal antibody WIC-79.3 which is L. major-specific and does not react with L. donovani. When another monoclonal, WIC-108.3, which cross-reacts with several Leishmania species, is used to affinity purify GL from L. donovani, the eluted material can partially protect genetically resistant mice against L. major. Thus, GL cross-reactions may in part underlie the known protective effects of crude L. donovani antigens against L. major infection. Experiments with live parasites of the L. major isolate LRC-L119, that is non-pathogenic in mice, that does not survive long in macrophages in vitro, and that has not been shown to contain any WIC-79.3 reactive GL, indicated that this isolate will very effectively protect mice against subsequent infection. This raises the possibility that GL is only one of at least two different classes of vaccinating antigen capable of protectively immunizing mice in this cutaneous leishmaniasis model.

Animals↗
PubMed1987

Responses in mice to Sj26, a glutathione S-transferase of Schistosoma japonicum worms.

The genetic variation in antibody responses of mice to glutathione S-transferase (GST) enzymes of Schistosoma japonicum worms, and in particular to a Mr 26,000 species termed Sj26, was analysed. Sera from infected mice, or mice immunized with adjuvant and an Sj26 beta-galactosidase fusion protein produced in Escherichia coli (Sj26FP), or purified near-native recombinant Sj26 produced in E. coli (rSj26), were assayed by enzyme-linked immunosorbent assay (ELISA) for antibody titres to GST purified from adult worms. Anti-GST antibody levels are high in a mouse strain, WEHI 129/J, that is genetically resistant to infection with S. japonicum. Antibody responses to GST are low in BALB/c mice and intermediate in most other mouse strains analysed such as CBA/H and C57B1/6. Responsiveness to Sj26 in adjuvant is dominant in (BALB/c x WEHI 129/J)F1 hybrids. BALB/c.H-2b and BALB/c.H-2k mice are higher responders than BALB/c. One feature of antibody responses to Sj26 is the variability within a group of mice. When rSj26 conjugated to the hapten azobenzenearsonate was used as immunogen, BALB/c mice produced substantial amounts of anti-Sj26 antibodies. In an attempt to correlate antibody levels with resistance in infected mice, a new functional assay was devised to measure the ability of sera to inhibit the binding of rSj26 to glutathione. However, there was no correlation between inhibitory titre in this assay and the numbers of worms recovered. In regard to the candidacy of GST as a vaccinating antigen in schistosomiasis japonica, the data raise the problem of variable responsiveness to the antigen that will need to be overcome by antigen modification and/or powerful adjuvants.

Animals↗
PubMed1987

Lipophosphoglycan of Leishmania major that vaccinates against cutaneous leishmaniasis contains an alkylglycerophosphoinositol lipid anchor.

The major cell surface glycoconjugate of Leishmania major, a putative parasite receptor for macrophages, is a lipophosphoglycan containing 81.6% (wt/wt) carbohydrate, 17.0% (wt/wt) phosphate, and 1.4% (wt/wt) lipid. It has been purified to homogeneity by hydrophobic chromatography and consists of a polydisperse family of molecules with Mr 5000-40,000. It contains galactose, mannose, glucose, arabinose, glucosamine, and inositol in the molar ratio of 51:21:5:6:1:1. The lipophosphoglycan has a complex structure, consisting mainly of tri- and tetrasaccharide units linked by phosphodiester bonds, which are cleaved by HF hydrolysis. The phosphate groups are located on the 6-hydroxyl of both galactose and mannose residues. The lipophosphoglycan is anchored to the parasite surface by a 1-O-alkyl-sn-glycero-3-phosphoinositol moiety. This conclusion is supported by analysis of the products of nitrous acid deamination, HF hydrolysis, and Staphylococcus aureus phosphatidylinositol specific-phospholipase C treatment. The 24:0 and 26:0 alkyl chains accounted for 93% of the ether-linked fatty acids in the lipid anchor. The results are also consistent with a glycosidic linkage between the inositol and a non-N-acetylated glucosamine residue. The lipophosphoglycan membrane anchor shares limited structural homology with the glycosylphosphatidylinositol anchors of several eukaryotic proteins, indicating that this type of membrane anchor is not limited to proteins. Vaccination of mice with the purified L. major lipophosphoglycan in liposomes induced resistance against cutaneous leishmaniasis.

Animals↗
PubMed1987

Evidence of anti-embryonation immunity and egg destruction in mice sensitized with immature eggs of Schistosoma japonicum.

BALB/c mice sensitized by repeated injections of immature eggs of the trematode worm, Schistosoma japonicum, were challenged with low numbers of cercariae and evidence was sought for inhibition of embryonation by examination of eggs in livers and intestines at days 40 - 42 of infection. In contrast to the situation in unsensitized control mice, a greater proportion of dead eggs was noted in tissues of many of egg-sensitized mice. There was also a decrease in the proportion of mature eggs relative to control mice. A substantial number of egg - sensitized mice contained no eggs in the liver though eggs were readily detected in their intestinal walls. The data support the concept that immune effector mechanisms act on eggs in a manner that prevents their full development into a miracidium and thus a rich source of immunopathologic antigens.

Animals↗
PubMed1987

Leishmania major: a very sensitive dot-blot ELISA for detection of parasites in cutaneous lesions.

There is a need for accurate, rapid and early diagnosis of leishmaniasis, which would distinguish between the benign and severe forms of the disease. We have used a monoclonal antibody directed to a polymorphic, species-specific antigen in diagnostic assays for leishmaniasis. The dot-blot enzyme-linked immunosorbent assay described here can detect as few as 300 culture promastigotes and 20,000 amastigotes of Leishmania major with no cross-reaction with other species and no background from skin macrophages or other cells. This level of sensitivity is sufficient to detect parasite antigen aspirated in a few microliters of liquid from small lesions in mice. This assay could form the basis for a sensitive, rapid and inexpensive dip-stick test for large-scale use for the diagnosis and epidemiology of cutaneous leishmaniasis.

Animals↗
PubMed1986

Passive transfer of Leishmania lipopolysaccharide confers parasite survival in macrophages.

Infection of macrophages by the intracellular protozoan parasite Leishmania involves specific attachment to the host membrane, followed by phagocytosis and intracellular survival and growth. Two parasite molecules have been implicated in the attachment event: Leishmania lipopolysaccharide (L-LPS) and a glycoprotein (gp63). This study was designed to clarify the role of L-LPS in infection and the stage in the process of infection at which it operates. We have recently identified a Leishmania major strain (LRC-L119) which lacks the L-LPS molecule and is not infective for hamsters or mice. This parasite was isolated from a gerbil in Kenya and was identified phenotypically as L. major by isoenzyme and fatty acid analysis. In this study we have confirmed at the genotype level that LRC-L119 is L. major by analyzing and comparing the organization of cloned DNA sequences in the genome of different strains of L. major. Here we show that LRC-L119 promastigotes are phagocytosed rapidly by macrophages in vitro, but in contrast to virulent strains of L. major, they are then killed over a period of 18 hr. In addition, we show that transfer of purified L-LPS from a virulent clone of L. major (V121) into LRC-L119 promastigotes confers on them the ability to survive in macrophages in vitro.

Animals↗
PubMed1986

Identification of a particular antigen from a parasite cDNA library using antibodies affinity purified from selected portions of Western blots.

Portions of nitrocellulose filters containing blotted electrophoresed antigens of Schistosoma japonicum adult worms were reacted with polyclonal rabbit antisera raised to this human parasite. Eluted antibodies were used as probes for detection of antigen-positive clones in an Escherichia coli lambda gt11 amp3 expression library of adult worm cDNA. Several cloned antigens corresponding to a S. japonicum antigen of Mr 26 000, being sought as a candidate vaccine molecule in a mouse model of schistosomiasis japonica, were identified using this approach. The method provides an antibody reagent that is an attractive alternative to other more tedious means of producing oligospecific antibodies, including monoclonal antibodies, for screening of expression libraries.

Animals↗
PubMed1986

Expression of Schistosoma japonicum antigens in Escherichia coli.

A cloned library of DNA complementary to the mRNA of adult Schistosoma japonicum has been prepared and expressed as fusion proteins with Escherichia coli beta-galactosidase. Colonies expressing the S. japonicum cDNA clones were screened both with antibodies from individuals with a history of schistosomiasis and with antibodies obtained from a rabbit immunized with whole adult worms. In both cases colonies were detected which bound antibody, although the frequency of antigen-positive clones was much higher with the rabbit antiserum than with human sera. In both cases the proportion of colonies reacting with antibodies was markedly lower than that published for equivalent screens of Plasmodium falciparum cDNA with sera from individuals with a history of falciparum malaria. Several major S. japonicum antigens were identified by the affinity purification of antibodies using immobilised fusion proteins produced during lytic growth of the recombinant bacteriophage.

Antibodies↗