Biomedical subjects

P Perlmann

Publications and source records attributed to P Perlmann.

At least 37 records · Page 2Linked to original sources

PubMed1997

A malariometric survey in a rural community in the Muheza district, Tanzania: age profiles in the development of humoral immune responses.

A malariometric survey was carried out in a rural community situated in a malaria holoendemic endemic area of Tanzania. A random sample (n = 228) of different age groups was taken to elucidate the association between anti-Pf155/RESA and anti-Pf332 antibody responses and classical malaria indices. Parasitaemia, fever, splenomegaly, haematocrit and antimalarial consumption were assessed. Antibody responses against Pf155/RESA and Pf332 peptides were determined by ELISA. The age profiles of parasite density, splenomegaly, fever, haematocrit values and prevalence of antibody responses indicated intensive malaria exposure and the highest impact of malaria in small children. Forty-five percent of the study population had detectable chloroquine and desethyl-chloroquine blood levels, and the highest frequency and concentrations were recorded in the 12-23 months old. There was no significant association between the presence of drug and parasite density in the different age groups, although in the < 15 years old there was lower parasite prevalence among the children positive for drug in their blood (P < 0.05). High prevalence of antibody responses to all antigens was observed already at an early age, but the mean anti-Pf155/RESA and anti-Pf332 antibody levels increased significantly only in the adult group (P < 0.01). Significantly lower mean parasite densities were observed in high responders to Pf155/RESA and Pf332 peptides for the > or = 10 years old. For the 1-9 years, a similar difference was only observed in the high responders to Pf332. For the whole material, anti-Pf155/RESA and anti-Pf332 antibody levels correlated positively with age. When the effect of age was allowed for in analysing the relationship between parasite density and antibody level against the different antigens, a significant negative correlation was found only with regard to Pf332 in the > = 10 years age group. These results suggest that anti-Pf332 antibodies appear to be a better indicator for antiparasitic immunity, but both antigens are important for immune protection.

Adolescent↗

PubMed1997

Predominance of H-2d- and H-2k-restricted T-cell epitopes in the highly repetitive Plasmodium falciparum antigen Pf332.

Genetic restriction of immune responses to malaria antigens is an important issue for a better comprehension of malaria immunity as well as for development of subunit vaccines. To experimentally define the major histocompatibility complex restriction of immune responses to the highly repetitive Plasmodium falciparum high-molecular-weight antigen Pf332, H-2-congenic mice were immunized with EB200, a recombinant fragment of Pf332 consisting of degenerate repeat motifs. Strong B- and T-cell responses were elicited in H-2d and H-2k mice whereas responses in H-2b, H-2q and H-2s mice were of lower magnitude. The T-cell specificity elicited by EB200 was defined by in vitro proliferative responses to a panel of overlapping peptides spanning EB200. Dominant epitopes were identified for H-2d and H-2k mice, respectively, and an additional epitope was recognized by all five mouse strains. Selected EB200-derived peptides were further investigated for their ability to elicit T-cell help when injected as multiple antigen peptides. Defined H-2d- and H-2k-restricted T-cell epitopes generated high antibody levels in the respective mouse strains, as did several peptides lacking defined epitopes indicating the presence of additional H-2d- and H-2k-restricted, cryptic or subdominant T-cell epitopes in EB200. The biased H-2 restriction pattern of T-cell epitopes in Pf332 and, as previously reported, in structurally related repeats in the malaria antigens Pf11.1 and Pf155/RESA may be explained by a shared motif for H-2d and H-2k class II-restricted T-cell epitopes, as revealed by alignment of these sequences.

Amino Acid Sequence↗

PubMed1997

Plasmodium falciparum: selection of parasite subpopulations with decreased sensitivity for antibody-mediated growth inhibition in vitro.

The chronic, recrudescent nature of malaria has been linked to antigenic diversity of the parasite in which protective immunity against Plasmodium falciparum may be obtained after repeated exposure to infection during a long time. In this study we show that parasite populations with decreased sensitivity to antibody-mediated growth inhibition are readily generated in vitro. A laboratory strain, F32, was cultured for long periods (10-12 weeks) in the presence of suboptimal inhibitory antibody concentrations. The antibodies used were the human monoclonal antibody 33G2 reacting with a linear epitope of the P. falciparum blood-stage antigen 332 and rabbit antibodies to repeat sequences of the blood-stage antigen Pf155/RESA. Our data indicate that the P. falciparum parasites adapt to antibody pressure as reflected by their specifically decreased sensitivity to growth inhibition. A relative resistance of the parasite to growth inhibition mediated by the antibodies used in the culture developed successively, while the parasite remained sensitive to growth inhibition by other antibodies. When the antibody pressure was removed a successive return of sensitivity to growth inhibition developed. Immunoflourescence did not reveal any significant difference in antigen expression between the parasite populations. However, PCR analysis showed that a new population appeared in the parasites grown in the presence of mAb 33G2, while no such change was detected in those grown in the presence of the rabbit antibodies. Our data suggest that the specific decrease in sensitivity to growth inhibition may either be due to down-regulation of antigen synthesis or expression by antibody pressure or, that antibody pressure selects for parasites with low expression of a specific antigen from a heterogeneous parasite population.

Animals↗

PubMed1997

Elevated plasma levels of IgE in Plasmodium falciparum-primed individuals reflect an increased ratio of IL-4 to interferon-gamma (IFN-gamma)-producing cells.

People living in Plasmodium falciparum-endemic areas frequently have elevated levels of total as well as P. falciparum-specific serum IgE. This study aimed at investigating whether the elevated serum IgE levels reflect a shift in the balance between CD4+ T helper 1 (Th1) and T helper 2 (Th2) cells in individuals naturally exposed to the P. falciparum parasite. To investigate the role of Th1 and Th2 cells in the human P. falciparum system we used the ELISPOT assay to determine the ratio of IFN-gamma- and IL-4-producing cells after specific antigen or mitogen activation in vitro. The donors were individuals who had acquired immunity through natural exposure to the parasite. In response to the specific malaria antigens, very few IL-4-producing cells were seen. However, in the response of individual donors to the polyclonal T cell activator, leucoagglutinin (La), the anti-malarial IgE levels in plasma were correlated with an increased ratio of IL-4/IFN-gamma producing cells. Thus, donors with ratios of IL-4/IFN-gamma > 1 exhibited mean plasma anti-malarial IgE levels significantly greater than those with ratios < 1. In individuals not living in P. falciparum-endemic areas the ratio of IL-4/IFN-gamma was always < 1. Taken together, our data suggest a shift in the balance between Th1 and Th2 cells in naturally P. falciparum-primed individuals, associated with elevated anti-P. falciparum plasma IgE levels. The role and biological significance of IgE (Th2-type immune response) for protection against P. falciparum and/or pathogenesis of malaria require further study.

Adolescent↗

PubMed1997

Wild isolates of Plasmodium falciparum malaria show decreased sensitivity to in vitro inhibition of parasite growth mediated by autologous host antibodies.

Antigenic diversity in field populations of Plasmodium falciparum parasites may delay the acquisition of protective immunity to malaria, the development of which may thus require repeated exposure to infection over a prolonged period of time. In this study we show that P. falciparum parasites may vary in their sensitivity to antibody-mediated invasion/growth inhibition in vitro. Wild isolates of P. falciparum from children living in an endemic area of Burkina Faso were tested for their sensitivity to the growth inhibitory effects of antibodies originating from the same (autologous) and from other donors (heterologous). A significantly lower invasion inhibition activity was obtained when the isolates and antibodies were tested in autologous compared with heterologous combinations. The lower sensitivity to growth inhibition by autologous antibodies may be due to immune pressure in vivo, selecting from a heterogeneous parasite population those with a low expression of the antigens recognized by the host's antibodies. Alternatively, the parasites cultured from each child might represent expanding parasite populations, mainly constituting strains not earlier seen by the immune system of that specific host. The results reinforce the concern about Plasmodium antigenic diversity as a major obstacle towards the development of an effective malaria vaccine.

ABO Blood-Group System↗

PubMed1997

Comparative study of DNA-based immunization vectors: effect of secretion signals on the antibody responses in mice.

The presence of a signal sequence preceding the gene encoding a target antigen in a DNA vaccine should facilitate secretion of the in vivo translated antigen. The immune responses elicited upon injection with such a vector could differ from those induced by the same vector lacking a signal sequence. In the present study, the humoral responses elicited in mice immunized with two plasmids, either containing or lacking the human tissue plasminogen activator signal sequence, were compared. Both plasmids encode the chimeric antigen ZZN4, containing a malaria antigen Pf332-derived sequence (N4) linked to a bacterial fusion partner (ZZ). In vitro transfection of COS cells with each plasmid and treatment of the transfectants with brefeldin A confirmed that secretion of ZZN4 via the endoplasmic reticulum and Golgi pathway only occurred in cells transfected with the signal peptide-encoding plasmid. Repeated intramuscular injections of mice with either of the plasmids elicited comparable antibody responses to ZZN4 with regard to kinetics, specific IgG levels and persistence. These results indicate that in vivo transfection of muscle cells by either of these two plasmids generated comparable levels of antigen available for B-cell recognition and for uptake by antigen-presenting cells, despite the differential intracellular targeting of the encoded antigen. The relevance of these findings for the design of DNA vaccine vectors is discussed.

Animals↗

PubMed1997

Immunoglobulin E, a pathogenic factor in Plasmodium falciparum malaria.

Most children and adults living in areas where the endemicity of Plasmodium falciparum malaria is high have significantly elevated levels of both total immunoglobulin E (IgE) and IgE antimalarial antibodies in blood. This elevation is highest in patients with cerebral malaria, suggesting a pathogenic role for this immunoglobulin isotype. In this study, we show that IgE elevation may also be seen in severe malaria without cerebral involvement and parallels an elevation of tumor necrosis factor alpha (TNF). IgE-containing serum from malaria immune donors was added to tissue culture plates coated with rabbit anti-human IgE antibodies or with P. falciparum antigen. IgE-anti-IgE complexes as well as antigen-binding IgE antibodies induced TNF release from peripheral blood mononuclear cells (PBMC). Nonmalaria control sera with no IgE elevation induced significantly less of this cytokine, and the TNF-inducing capacity of malaria sera was also strongly reduced by passing them over anti-IgE Sepharose columns. The cells giving rise to TNF were adherent PBMC. The release of this cytokine probably reflects cross-linking of their low-affinity receptors for IgE (CD23) by IgE-containing immune complexes known to give rise to monocyte activation via the NO transduction pathway. In line with this, adherent monocytic cells exposed to IgE complexes displayed increased expression of CD23. As the malaria sera contained IgG anti-IgE antibodies, such complexes probably also play a role in the induction of TNF in vivo. Overproduction of TNF is considered a major pathogenic mechanism responsible for fever and tissue lesions in P. falciparum malaria. This overproduction is generally assumed to reflect a direct stimulation of effector cells by certain parasite-derived toxins. Our results suggest that IgE elevation constitutes yet another important mechanism involved in excessive TNF induction in this disease.

Adult↗

PubMed1996

T cell-derived IL-3 induces the production of IL-4 by non-B, non-T cells to amplify the Th2-cytokine response to a non-parasite antigen in Schistosoma mansoni-infected mice.

We describe a novel amplification mechanism underlying the increased early IL-4 production observed in Schistosoma mansoni-infected mice in response to a non-parasite Ag, sperm whale myoglobin (SwMb). Earlier studies have shown that splenic Fc epsilon R+ non-B, non-T (NBNT) cells from schistosome-infected mice secrete IL-4 after stimulation with parasite Ag. We now demonstrate that purified NBNT cells from SwMb-immunized S. mansoni-infected mice do not respond directly to SwMb, but produce IL-4 in response to IL-3. Accordingly, we show that the early SwMb-specific IL-4 response of spleen cells (SC) from immunized infected mice is dependent on IL-3 and on CD4+ T cells. Thus, most of the early SwMb-induced IL-4 from SC of infected mice appears to be produced by NBNT cells triggered by IL-3 synthesized by SwMb-specific CD4+ T cells. IL-3-induced IL-4 production was also observed in purified NBNT cells from immunized uninfected mice, but the frequency and/or IL-4-producing capacity of splenic IL-3-responsive cells was found to be 8 to 16 times higher in immunized infected animals. IL-4 production by purified CD4+ cells from immunized infected mice was also seen after SwMb stimulation, but this response showed slower kinetics than those of total SC, was IL-3-independent, and on average threefold greater than that by CD4+ cells from immunized uninfected controls. Thus, increased SwMb-induced IL-4 production in immunized S. mansoni-infected mice results from direct synthesis by CD4+ T cells, as well as their stimulation via IL-3 of an expanded population of NBNT cells. The latter pathway may serve as an amplification loop for Th2-cytokine responses.

Animals↗

PubMed1996

Plasmodium falciparum: differential parasite growth inhibition mediated by antibodies to the antigens Pf332 and Pf155/RESA.

Plasmodium falciparum-reactive antibodies can inhibit growth of parasite blood stages in vitro by interfering at different stages of parasite development. Antibodies reactive with repeat sequences in Pf155/ring-infected surface antigen (Pf155/RESA) or in antigen Pf332 inhibit parasite growth as determined by a reduction of newly infected erythrocytes. Antibodies to Pf155/RESA have been implicated in merozoite invasion inhibition. Since it was considered unlikely that antibodies to the late stage antigen Pf332 act on merozoite invasion, we investigated at what developmental stage this growth inhibition takes place. Parasites were cultured in the presence of antibodies to repeat sequences of either Pf332 or Pf155/RESA and were examined with regard to the distribution of different stages and their morphology. As expected, antibodies to both antigens decreased the number of ring stages after reinfection. They did not affect the intraerythrocytic development of ring stages or trophozoites. However, antibodies to Pf332 induced high levels of schizonts displaying an abnormal morphology while antibodies to Pf155/RESA induced considerably lower levels of degenerated schizonts. Thus, Pf332-reactive antibodies seem to interfere with the schizont development by blocking the rupture of mature schizonts or, alternatively, by interfering intraerythrocytically with late stage parasites.

Amino Acid Sequence↗

PubMed1996

Lack of association between levels of transplacentally acquired Plasmodium falciparum-specific antibodies and age of onset of clinical malaria in infants in a malaria endemic area of Nigeria.

A cohort of 117 newborns was followed longitudinally for 12 months to determine the age of onset of clinical malaria and the subsequent episodes of malaria, and to investigate the possible existence of a correlation between level of transplacentally acquired Plasmodium falciparum-specific antibodies and age of onset of malaria in the infant. The mean age of onset of malaria in 49 infants was 4.48 +/- 1.54 months. Mean (+/- S.D.) age of onset of clinical malaria in haemoglobin AA infants (4.38 +/- 1.14) was significantly (P < 0.05) lower compared with haemoglobin AS (5.58 +/- 2.43) infants. No correlation was obtained between the age of onset of malaria and the level of cord serum total IgG, IgM and antibodies to P. falciparum antigens. Cord blood seropositivity for antibodies to the blood stage antigen Pf155/RESA and its C-terminal repeat sequence (EENV)6 or to the (NANP)6 peptide representing repeats of the circumsporozoite protein (CSP) did not influence the age of onset of clinical malaria. However, infants with haemoglobin AS whose cord blood was seropositive for antibodies to the (EENV)6 or (NANP)6 peptide showed delayed onset (P < 0.001) of malaria compared with AA seropositive infants. Although our results indicate that transplacentally acquired antibodies to the studied antigens alone offer no significant protection against malaria during the first few months of life, antibodies in concert with other factors such as haemoglobin genotype may contribute to the protection of the newborn.

Age Factors↗

PubMed1996

Immune responses in congenic mice to multiple antigen peptides based on defined epitopes from the malaria antigen Pf332.

Repeat sequences from the Plasmodium falciparum blood stage antigen Pf332 frequently comprise the pentapeptide VTEEI, an epitope recognized by certain parasite neutralizing antibodies. This B-cell epitope was assembled in an octavalent multiple antigen peptide (MAP) system either as trimers (VTEEI)3 (MAP1) or as an integral part of a naturally occurring Pf332 undecamer repeat sequence SVTEEIAEEDK (MAP2). Characteristics of the immunogenicity of these subunit constructs were evaluated in H-2 congenic mice. MAP1 generated antibody responses in mice of the H-2d, H-2k and H-2q haplotypes, but not in H-2b or H-2s mice, whereas MAP2 only induced antibodies in mice of H-2k haplotype. When analysing T-cell responses induced by the MAP, lymph node cells from responder strains primed in vivo with MAP1 proliferated in response to restimulation with both MAP1 and the peptide (VTEEI)3. MAP2, however, did not induce a detectable T-cell proliferation. Additionally, the lack of antibody response to MAP1 in H-2b mice could be circumvented by combining the MAP1 peptide and a H-2b-restricted T-cell epitope in a diepitope MAP construct. Despite the fact that the motif VTEEI has not been identified in Pf332 sequences in the form of a trimer, MAP1 did induce Pf332 protein-reactive antibodies. Assembly of multimers of short defined epitopes in MAP constitutes an interesting approach for the design of polyvalent subunit immunogens.

Amino Acid Sequence↗

PubMed1996

Immunoglobulin E elevation in Plasmodium chabaudi malaria.

In order to investigate the mechanism of immunoglobulin E (IgE) elevation in malaria we studies mice infected with asexual blood stages of the rodent malaria parasite Plasmodium chabaudi chabaudi for total IgE and IgE antimalarial antibodies. Multiply infected mice had elevated levels of total as well as malaria-specific IgE in their sera. Sera taken from mice 3 weeks after one infection with P. chabaudi showed no IgE elevation, indicated that prolonged or repeated exposure to the parasite is necessary for the induction of an IgE response, which also is induced independently of previous or simultaneous infection with other pathogens such as helminths.

Animals↗

PubMed1996

Generation of in vitro natural cytotoxicity of horse lymphocytes against sarcoid-derived tumor cells not expressing major histocompatibility complex antigens.

OBJECTIVE: To analyze in vitro lymphocyte-mediated immune responses of horses with sarcoids against allogeneic sarcoid cells containing endogenous retrovirus but not expressing major histocompatibility complex antigens. DESIGN: Lymphocyte-mediated immune reactions were assessed by means of proliferative responses in mixed lymphocyte tumor cell culture (MLTC) assay and lymphocyte-mediated cytotoxicity against various equine target cells. ANIMALS: 12 horses with sarcoid tumors and 15 control horses. PROCEDURE: Blood lymphocytes were cocultured in MLTC with allogeneic sarcoid cells (Mc-1, BayMc-1), equine testis cells, or normal equine dermal fibroblasts. Lymphocytes were assayed for proliferative responses by [3H]thymidine uptake and for cytotoxicity against the same targets by 51Cr release assay. The lymphocyte populations were analyzed for some common surface markers. RESULTS: Lymphocytes from horses with sarcoids exerted an anamnestic proliferative response in MLTC against Mc-1 cells, but this procedure never generated cytotoxic lymphocytes. However, lymphocytes from all horses cultured in medium with 10% allogeneic serum only had selective. natural cytotoxicity against Mc-1 that was generated without DNA synthesis. Approximately 80% of the lymphocytes disappeared during culture; however the remaining population of small, viable lymphocytes indicated a decrease of CD4+ T lymphocytes, but numbers of T cells with receptors for Helix pomatia A hemagglutinin were unaffected. Few lymphocytes had Fc-receptors for IgG, were complement-reactive positive cells or were B cells expressing surface immunoglobulin. CONCLUSIONS: Results may indicate a natural defense system, which preferentially recognizes and lyses tumor cells that are deficient in surface expression of major histocompatibility complex antigens, without intervention of conventional T-cell receptors or antibodies.

Animals↗

PubMed1995

A longitudinal study of seroreactivities to Plasmodium falciparum antigens in Nigerian infants during their first year of life.

The kinetics of passively transferred maternal antibodies to antigens of Plasmodium falciparum and the dynamics of acquisition of these antibodies during the first year of life was investigated in infants born in a malaria endemic area of south-western Nigeria. Blood samples were collected from the infants at bi-monthly follow-up visits for the analysis of total serum immunoglobulin G, IgM, IgA and antibodies to the antigen Pf155/RESA and against synthetic peptides representing antigenic sequences of the blood stage antigen Pf155/RESA and Ag332 or the circumsporozoite protein (CSP). IgG levels fell from birth till 4 months and a steady rise was observed thereafter till ten months of life. On the contrary mean IgM and IgA levels increased throughout the first year of life. Generally the number of infants positive for antibodies to the antigens under investigation fell from birth and between 4-6 months of age was either low or absent. None of the infants were positive for antibodies to the peptide representing Ag332 during the first year of life. The earliest seroconversion was detected at 6 months of age involving the Pf155/RESA and (NANP)6 antigens. The results indicate a high level of exposure in this study area to malaria infection early in life. The finding of an active antibody response to malarial antigens in infancy encourages the hope that a malaria vaccine administered early in life may accelerate the development of naturally acquired immunity and thus protect the population most at risk.

Amino Acid Sequence↗

PubMed1995

Cytoadherence-related homologous motifs in Plasmodium falciparum antigen Pf155/RESA and erythrocyte band 3 protein.

Cytoadherence of Plasmodium falciparum-infected erythrocytes plays an important role in the pathogenesis of cerebral malaria. The identity of cell surface molecules on parasitized erythrocytes involved in cytoadherence is of great interest to understand the molecular basis of this mechanism. Peptide sequences derived from exofacial loops of the erythrocyte antigen band 3 from parasitized erythrocytes have previously been shown to inhibit cytoadherence. We now report that a non-repeated region of Pf155/RESA (residues 213-218) contains a hexapeptide motif being highly homologous to cytoadherence inhibitory sequences from band 3. Synthetic peptides containing the hexapeptide motif of Pf155/RESA inhibited the binding of P. falciparum-infected erythrocytes to melanoma cells in vitro. Furthermore, individuals residing in malaria-endemic areas have antibodies reactive with epitopes involving these motifs in band 3 and in Pf155/RESA.

Amino Acid Sequence↗

PubMed1995

Antibodies to Pf155/RESA and circumsporozoite protein of Plasmodium falciparum in paired maternal-cord sera from Nigeria.

Paired maternal-cord serum samples were analysed for antibodies to the Pf155/RESA and circumsporozoite protein (CSP) antigens of Plasmodium falciparum. Malaria parasites were found in 2.6% (3/117) of cord blood and 22.4% (26/116) of maternal samples. Immunofluorescence assays detected P. falciparum-specific IgG antibodies in all paired samples while P. falciparum-specific IgM was detected in 5.8% (7/121) of cord samples. The positivity rates for antibodies to Pf155/RESA and (NANP)6 but not (EENV)6, a C-terminal repeat sequence of Pf155/RESA, were significantly higher in maternal as compared with cord samples. Seropositivity rates to Pf155/RESA and (EENV)6 were not related to maternal parity group while positivity rates to the (NANP)6 peptide were higher in primiparae and multiparae of > or = 4 parity. These data confirm the transplacental transfer of P. falciparum-specific antibodies and the higher incidence of malaria parasitaemia in primiparae. The presence of P. falciparum-specific IgM in some cord samples suggests intrauterine sensitization of the foetus to malarial antigens.

Adolescent↗

PubMed1995

Immunogens containing sequences from antigen Pf332 induce Plasmodium falciparum-reactive antibodies which inhibit parasite growth but not cytoadherence.

Immunogens based upon sequences from the P. falciparum asexual blood stage antigen Pf332 were assessed for their capacity to induce antibodies inhibiting parasite growth or cytoadherence of infected erythrocytes in vitro. Selection of the Pf332 sequences was based on their reactivity with the human monoclonal antibody (MoAb) 33G2 which inhibits parasite growth as well as cytoadherence in vitro. Octameric multiple antigen peptides (MAP) were assembled based upon either a trimer of the minimal epitope recognized by the MoAb, VTEEI, or a Pf332 sequence including that motif, SVTEEIAEEDK. A dimer of SVTEEIAEEDK was also expressed in Escherichia coli, genetically fused to ZZ, two IgG-binding domains of staphylococcal protein A. Rabbit antibodies elicited by the immunogens reacted with Pf332 in immunofluorescence and in ELISA with Pf332 peptides which were also recognized by MoAb 33G2. The MAP with branched (VTEEI)3 peptide induced the highest titres of P. falciparum-reactive antibodies. In contrast to MoAB 33G2, none of the polyclonal Pf332 reactive sera cross-reacted with repeat sequences of the malaria antigen Pf155/RESA. The polyclonal Pf332-reactive antibodies inhibited parasite growth efficiently but had no or very low inhibitory effect in a cytoadherence assay. Thus, while Pf332 may be an important target for parasite neutralizing antibodies its involvement in cytoadherence is unclear.

Amino Acid Sequence↗

PubMed1995

Surface display compared to periplasmic expression of a malarial antigen in Salmonella typhimurium and its implications for immunogenicity.

Two different expression systems were investigated for the production of an 80 amino acid polypeptide, M3, from the C-terminus of the Plasmodium falciparum blood stage antigen Pf155/RESA in an attenuated Salmonella typhimurium vaccine strain. Upon expression, the malarial polypeptide was targeted either to the periplasm as a soluble fusion protein containing two IgG-binding domains (ZZ) from the staphylococcal protein A or, to the bacterial surface as an insert within a chimeric outer membrane protein A (OmpA) derived from Escherichia coli and Shigella dysenteriae. Both the ZZM3 and the OmpAM3 proteins were stably expressed in the periplasm or on the surface of Salmonella, respectively. The ZZ expression system yielded 10-100 times more malarial immunogen than did the OmpA system. Live recombinant Salmonella expressing ZZM3 or OmpAM3 were used to immunize mice intraperitoneally. Both the ZZM3 and OmpAM3 genes persisted for up to three weeks in bacteria isolated from different lymphoid organs. Bacteria expressing ZZM3 induced antibodies to M3, ZZ and to the Pf155/RESA antigen whereas, bacteria producing OmpAM3 induced similar levels of antibodies reactive with M3 but not with Pf155/RESA. Both recombinants induced a memory response of antibodies reactive with both M3 and Pf155/RESA. The high levels of M3 produced by the ZZ expression system make it suitable for the expression of heterologous antigens in Salmonella. Nevertheless, in spite of the quantitative difference in M3 expression, the ZZ and OmpA constructs elicited comparable immune responses to M3.

Animals↗