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P Druilhe

Publications and source records attributed to P Druilhe.

At least 19 recordsLinked to original sources
PubMed2004

Improved quantification of Plasmodium exoerythrocytic forms in rodents.

The result of a Plasmodium sporozoite challenge is currently evaluated either by detecting the emergence or not of parasites in the blood, or by estimating the "prepatent period", which is the time between sporozoite inoculation and the appearance of parasites in the blood. This type of measurement is relatively rough and has given way to another method of measuring sporozoite infectivity, which is to enumerate the exoerythrocytic forms (EEF) by microscopic examination of liver sections. Up until now, two different methods have been proposed to calculate and estimate the number of Plasmodium EEF forms in the livers of infected rodents, both of which are unfortunately biased to some extent. Here, we propose a different method of calculation, which more faithfully reflects the EEF number in the liver. This method is based on the calculated mean number of consecutive liver sections in which a schizont appears, and is host related.

Animals↗
PubMed2004

Effect on antibody and T-cell responses of mixing five GMP-produced DNA plasmids and administration with plasmid expressing GM-CSF.

One potential benefit of DNA vaccines is the capacity to elicit antibody and T-cell responses against multiple antigens at the same time by mixing plasmids expressing different proteins. A possible negative effect of such mixing is interference among plasmids regarding immunogenicity. In preparation for a clinical trial, we assessed the immunogenicity of GMP-produced plasmids encoding five Plasmodium falciparum proteins, PfCSP, PfSSP2, PfEXP1, PfLSA1, and PfLSA3, given as a mixture, or alone. The mixture induced higher levels of antibodies against whole parasites than did the individual plasmids, but was associated with a decrease in antibodies to individual P. falciparum proteins. T-cell responses were in general decreased by administration of the mixture. Immune responses to individual plasmids and mixtures were generally higher in inbred mice than in outbreds. In inbred BALB/c and C57BL/6 mice, coadministration of a plasmid expressing murine granulocyte-macrophage colony-stimulating factor (mGM-CSF), increased antibody and T-cell responses, but in outbred CD-1 mice, coadministration of mGM-CSF was associated with a decrease in antibody responses. Such variability in data from studies in different strains of mice underscores the importance of genetic background on immune response and carefully considering the goals of any preclinical studies of vaccine mixtures planned for human trials.

Animals↗
PubMed2004

Immunization of Saimiri sciureus monkeys with Plasmodium falciparum merozoite surface protein-3 and glutamate-rich protein suggests that protection is related to antibody levels.

The immunogenicity and protective efficacy of various antigen-adjuvant formulations derived either from the merozoite-surface protein-3 (MSP-3) or the glutamate-rich protein (GLURP) of Plasmodium falciparum were evaluated in Saimiri sciureus monkeys. These proteins were selected for immunogenicity studies based primarily on their capacity of inducing an antibody-dependent cellular inhibition effect on parasite growth. Some of the S. sciureus monkeys immunized with MSP-3(212-380)-AS02 or GLURP(27-500)-alum were able to fully or partially control parasitaemia upon an experimental P. falciparum [Falciparum Uganda Palo Alto (FUP-SP) strain] blood-stage infection, and this protection was related to the prechallenge antibody titres induced. The data are indicative that MSP-3 and GLURP can induce protective immunity against an experimental P. falciparum infection using adjuvants that are acceptable for human use and this should trigger further studies with those new antigens.

Animals↗
PubMed2003

[Cohorts and bio-libraries for studying malaria in tropical areas].

The relationship between Plasmodii transmission, infection, morbidity, genetic susceptibility and acquisition of natural immunity is studied among two cohorts in the Senegalese villages of Dielmo (300 inhabitants) and Ndiop (350 inhabitants) where malaria is holoendemic (about 200 P. falciparum infective bites/person/year) and mesoendemic (about 20 P. falciparum infective bites/person/year), respectively. The populations are under a daily active clinical survey. Blood samples are collected at least once per month. Plasma and red blood cells are stored in bio-libraries that allow longitudinal studies of the immune responses against plasmodial antigens and the investigation of the natural history of P. falciparum infections by molecular genotyping methods.

Animals↗
PubMed2001

Expression of the erythrocyte-binding antigen 175 in sporozoites and in liver stages of Plasmodium falciparum.

Screening of a Plasmodium falciparum genomic expression library for antigens expressed at the pre-erythrocytic stages resulted in the isolation of a recombinant phage (DG249) whose insert corresponded to regions II and III of a 175-kDa erythrocyte-binding antigen (EBA-175). EBA-175 is a parasite ligand implicated in red blood cell invasion. Reverse-transcriptase polymerase chain reaction, indirect immunofluorescent antibody test, and Western blot analysis confirmed that EBA-175 is expressed not only in blood-stage parasites but also in infected hepatocytes and on the sporozoite surface. The presence of EBA-175 on pre-erythrocytic parasites enhances the vaccine potential of this antigen by adding another target to the immune responses elicited by immunization.

Animals↗
PubMed2001

Long synthetic peptides encompassing the Plasmodium falciparum LSA3 are the target of human B and T cells and are potent inducers of B helper, T helper and cytolytic T cell responses in mice.

We synthesized 17 long synthetic peptides (LSP) spanning the whole 200-kDa Plasmodium falciparum liver stage antigen-3 (LSA3), an antigen that induces protection in chimpanzee, and analyzed their immunogenicity in BALB/c mice and their antigenicity in individuals living in a hyper-endemic malaria area. Our findings show that both specific antibodies and T cell proliferation against most LSA3-LSP develop in malaria-exposed adults. All individuals studied had detectable antibodies against a minimum of 6 and a maximum of 15 polypeptides. It is noteworthy that antibody prevalence and titers were as high against non-repeat as repeat regions. Although the extent of T cell reactivity was lower than that observed for B cells, most of the sequences contained at least one T helper epitope, indicating that the majority of LSA3-LSP contain both B and T cell epitopes within the same sequence. Injection of LSA3-LSP with SBSA2 adjuvant in mice, showed strong immunogenicity for most of them, eliciting both T cell responses and specific antibody production. While all the peptides were immunogenic for B cells, different patterns of T cell responses were induced. These peptides were thus classified in three sets according to the levels of the T cell proliferative and of the IFN-gamma-specific responses. Importantly, antibodies and T cells against some of the LSP were able to recognize LSA3 native protein on P. falciparum sporozoites. Additionally, some LSP (44-119, 1026-1095, 1601-1712) also contained epitopes recognized by H-2(d) class I-restricted T cells. These results led to the identification of numerous domains that are highly antigenic and immunogenic within the LSA3 protein, and underline the value of the LSP approach for vaccine development.

Adult↗
PubMed2001

The Plasmodium falciparum knob-associated PfEMP3 antigen is also expressed at pre-erythrocytic stages and induces antibodies which inhibit sporozoite invasion.

The expression of the pfemp3 gene and the corresponding PfEMP3 knob-associated protein in the pre-erythrocytic stages of Plasmodium falciparum was demonstrated by RT-PCR, Western blots, IFAT and IEM. The antigen was found on the surface of the sporozoite and in the cytoplasm of mature hepatic stage parasites. Immunological cross-reactivity was observed with sporozoites from the rodent malaria parasites Plasmodium yoelii yoelii and Plasmodium berghei and was exploited to assess a potential role of this protein at the pre-erythrocytic stages. Specific antibodies from immune individuals were found to inhibit P. yoelii yoelii and P. berghei sporozoite invasion of primary hepatocyte cultures. PfEMP3 should now be added to the small list of proteins expressed at the pre-erythrocytic stages of P. falciparum, and its vaccine potential now deserves to be investigated.

Animals↗
PubMed2001

Human malaria in immunocompromised mice: new in vivo model for chemotherapy studies.

We have recently designed a new Plasmodium falciparum mouse model and documented its potential for the study of immune effector mechanisms. In order to determine its value for drug studies, we evaluated its response to existing antimalarial drugs compared to that observed in humans. Immunocompromised BXN (bg/bg xid/xid nu/nu) mice were infected with either the sensitive NF54 strain or the multiresistant T24 strain and then treated with chloroquine, quinine, mefloquine, or dihydroartemisinin. A parallelism was observed between previously reported human responses and P. falciparum-parasitized human red blood cell (huRBC)--BXN mouse responses to classical antimalarial drugs, measured in terms of speed of decrease in parasitemia and of morphological alterations of the parasites. Mice infected with the sensitive strain were successfully cured after treatment with either chloroquine or mefloquine. In contrast, mice infected with the multiresistant strain failed to be cured by chloroquine or quinine but thereafter responded to dihydroartemisinin treatment. The speed of parasite clearance and the morphological alterations induced differed for each drug and matched previously reported observations, hence stressing the relevance of the model. These data thus suggest that P. falciparum-huRBC-BXN mice can provide a valuable in vivo system and should be included in the short list of animals that can be used for the evaluation of P. falciparum responses to drugs.

Animals↗
PubMed2001

DNA immunization by Plasmodium falciparum liver-stage antigen 3 induces protection against Plasmodium yoelii sporozoite challenge.

DNA-based immunization of mice by Plasmodium falciparum liver-stage antigen 3 (PfLSA3), a novel highly conserved P. falciparum preerythrocytic antigen, was evaluated. Animals developed a dominant Th1 immune response (high gamma interferon T-cell responses and predominance of immunoglobulin G2a) to each of three recombinant proteins spanning the molecule. We have exploited the immunological cross-reactivity of PfLSA3 with its putative homologue on sporozoites of the rodent parasite Plasmodium yoelii, and we show for the first time that responses induced by PfLSA3 in mice significantly protect against a heterologous challenge by P. yoelii sporozoites. These results support a significant effect of DNA-induced immune responses on preerythrocytic stages.

Animals↗
PubMed2001

Human antibodies against Plasmodium falciparum liver-stage antigen 3 cross-react with Plasmodium yoelii preerythrocytic-stage epitopes and inhibit sporozoite invasion in vitro and in vivo.

The Plasmodium falciparum liver-stage antigen 3 (LSA3), a recently identified preerythrocytic antigen, induces protection against malaria in chimpanzees. Using antibodies from individuals with hyperimmunity to malaria affinity purified on recombinant or synthetic polypeptides of LSA3, we identified four non-cross-reactive B-cell epitopes in Plasmodium yoelii preerythrocytic stages. On sporozoites the P. yoelii protein detected has a molecular mass similar to that of LSA3. T-cell epitopes cross-reacting with P. yoelii were also demonstrated using peripheral blood lymphocytes from LSA3-immunized chimpanzees. In contrast, no cross-reactive epitopes were found in Plasmodium berghei. LSA3-specific human antibodies exerted up to 100% inhibition of in vitro invasion of P. yoelii sporozoites into mouse hepatocytes. This strong in vitro activity was reproduced in vivo by passive transfer of LSA3 antibodies. These results indicate that the homologous epitopes may be biologically functional and suggest that P. yoelii could be used as a model to assess the antisporozoite activity of anti-LSA3 antibodies.

Animals↗
PubMed2001

Selection of glutamate-rich protein long synthetic peptides for vaccine development: antigenicity and relationship with clinical protection and immunogenicity.

Antibodies against three long synthetic peptides (LSPs) derived from the glutamate-rich protein (GLURP) of Plasmodium falciparum were analyzed in three cohorts from Liberia, Ghana, and Senegal. Two overlapping LSPs, LR67 and LR68, are derived from the relatively conserved N-terminal nonrepeat region (R0), and the third, LR70, is derived from the R2 repeat region. A high prevalence of antibody responses to each LSP was observed in all three areas of endemic infection. Levels of cytophilic immunoglobulin G (IgG) antibodies against both GLURP regions were significantly correlated with protection from clinical P. falciparum malaria. Protected children from the Ghana cohort possessed predominantly IgG1 antibodies against the nonrepeat epitope and IgG3 antibodies against the repeat epitope. T-cell proliferation responses, studied in the cohort from Senegal, revealed that T-helper-cell epitopes were confined to the nonrepeat region. When used as immunogens, the LR67 and LR68 peptides elicited strong IgG responses in outbred mice and LR67 also induced antibodies in mice of different H-2 haplotypes, confirming the presence of T-helper-cell epitopes in these constructs. Mouse antipeptide antisera recognized parasite proteins as determined by immunofluorescence and immunoblotting. This indicates that synthetic peptides derived from relatively conserved epitopes of GLURP might serve as useful immunogens for vaccination against P. falciparum malaria.

Adolescent↗
PubMed2001

[Evaluation of in vitro drug sensitivity of antimalarials for Plasmodium falciparum using a colorimetric assay (DELI-microtest)].

The DELI-microtest (Double-Site Enzyme Linked LDH Immunodetection) is a colorimetric method for determination of Plasmodium falciparum in vitro sensitivity to antimalarial drugs. This method is based on the capture of the specific lactate deshydrogenase of Plasmodium falciparum (pLDH) using 2 monoclonal antibodies which recognize 2 different sites of the enzyme. The second monoclonal antibody is biotinylated and may catch a peroxidase-labelled streptavidine and develop a colorimetric reaction in the presence of peroxidase substrate. The level of pLDH released by parasite growth is determined by the measure of optical densities. An excellent correlation has been observed between the IC50's obtained using the DELI-microtest and the classical isotopic microtest, and the high sensitivity of the DELI-microtest can be used to measure the IC50's for isolates with parasitemia as low as 0.005%. It is easier and faster to process than isotopic microtest in field conditions. In addition, due to high sensitivity of the DELI-microtest, isolates with a low parasitemia may be included in the studies.

Animals↗
PubMed2001

Rodent malaria in the natural host--irradiated sporozoites of Plasmodium berghei induce liver-stage specific immune responses in the natural host Grammomys surdaster and protect immunized Grammomys against P. berghei sporozoite challenge.

The choice of the host in studying host-parasite interactions is of crucial importance, and the use of a natural host is most appropriate in answering pertinent questions related to human malaria. The Grammomys surdaster is the natural host and reservoir of the rodent malaria parasite Plasmodium berghei. This natural host is difficult to protect by irradiated sporozoite immunization, a situation comparable to what has been observed in humans with P. falciparum. This is in contrast to the complete protection that can be induced in artificial hosts like inbred mice strains. The natural host is highly susceptible to P. berghei hepatic stage infections. Immunization with irradiated sporozoites in Grammomys generates blocked hepatic stage parasites and immunized Grammomys protected upon live sporozoite challenge generate antibody and T cell proliferative responses to these hepatic stages. Associated with proliferation, cytokines are secreted into culture supernatants constituted mainly of Interferon gamma, negligible amounts of TNF-alpha, and no IL-4. Natural host-parasite interactions of Grammomys surdaster-P. berghei can help define the effector mechanism(s) in the Plasmodium falciparum-human interaction.

Animals↗
PubMed2000

Human malaria in immunocompromised mice: an in vivo model to study defense mechanisms against Plasmodium falciparum.

We have recently described that sustained Plasmodium falciparum growth could be obtained in immunodeficient mice. We now report the potential of this new mouse model by assaying the effect of the passive transfer of antibodies (Abs) which in humans have had a well-established effect.Our results show that the total African adult hyperimmune immunoglobulin Gs (HI-IgGs) strongly reduce P. falciparum parasitemia similarly to that reported in humans, but only when mice are concomitantly reconstituted with human monocytes (HuMNs). In contrast, neither HI-IgGs nor HuMNs alone had any direct effect upon parasitemia. We assessed the in vivo effect of epitope-specific human Abs affinity-purified on peptides derived either from the ring erythrocyte surface antigen (RESA) or the merozoite surface protein 3 (MSP3). The inoculation of low concentrations of anti-synthetic peptide from MSP3, but not of anti-RESA Abs, consistently suppressed P. falciparum in the presence of HuMNs. Parasitemia decrease was stronger and faster than that observed using HI-IgGs and as fast as that induced by chloroquine. Our observations demonstrate that this mouse model is of great value to evaluate the protective effect of different Abs with distinct specificity in the same animal, a step hardly accessible and therefore never performed before in humans.

Adult↗
PubMed2000

Identification of a major B-cell epitope of the Plasmodium falciparum glutamate-rich protein (GLURP), targeted by human antibodies mediating parasite killing.

The antigenicity of the glutamate-rich protein (GLURP) of Plasmodium falciparum was comprehensively evaluated in epitope-mapping studies utilizing a phage display library, synthetic peptides and anti-GLURP IgG preparations previously shown to promote strong antibody-dependent cellular inhibition (ADCI) effects. We identified six major B-cell epitopes within the nonrepetitive region R0, corresponding to amino acid residues 173 to 187 (P1), 193 to 207 (P3), 216 to 229 (P4), 264 to 288 (P11), 343 to 357 (P10), and 407 to 434 (S3). Of these, four (P1, P3, P4, and S3) were frequently recognized by high-titered IgG antibodies in plasma samples from immune Liberian adults (prevalence: 29.1-45.0%). The three epitopes P1, P3, and P4 contained a common motif (seven out of nine positions are identical) and may thus constitute a family of structurally related epitopes. This leaves two distinct epitopes, one (P3) representing this new epitope family and S3 as targets for biologically active antibodies. Human IgG antibodies from single plasma samples were affinity-purified against these peptides. P3-specific IgG preparations were consistently more effective in ADCI than S3-specific IgG. Among the different GLURP epitopes, we therefore suggest that the P3 epitope is potentially the most important epitope in GLURP for the development of clinical immunity to malaria in man.

Adult↗
PubMed2000

High immunogenicity in chimpanzees of peptides and lipopeptides derived from four new Plasmodium falciparum pre-erythrocytic molecules.

We have investigated the immunogenicity in chimpanzees of twelve synthetic peptides derived from four new Plasmodium falciparum molecules expressed at pre-erythrocytic stages of the human malaria parasite. These parasite molecules were initially selected through their ability to be recognized by stage restricted human antibodies. Twelve 20- to 41-mer peptides representing potential human B- or T-cell epitopes were selected from these proteins, and synthesized. Six of these were modified by a C-terminal lipidic chain in order to re-inforce their immunogenicity. Strong B- and T-helper cell responses were induced in chimpanzees by lipopeptides injected without adjuvant and by peptides in Montanide. All twelve peptides induced CD4(+) T-cell proliferative responses, as well as the secretion of IFN-gamma (some of them at very high levels) and eleven peptides induced antibody responses. The immune responses elicited in this way were reactive with native parasite proteins, as shown by recall studies with sporozoite stage proteins, and proved to be long-lasting (up to 10 months after immunization). Our results support the strategy employed to select these four new malarial antigens and the corresponding peptides, and suggest that the immunizing formulations are both efficient and clinically acceptable.

Amino Acid Sequence↗
PubMed2000

Plasmodium falciparum: a comparative analysis of the genetic diversity in malaria-mesoendemic areas of Brazil and Madagascar.

For a better definition of the polymorphic features of Plasmodium falciparum parasite populations, the polymerase chain reaction (PCR) typing technique was used to investigate the genetic diversity and complexity of parasites harbored by acute P. falciparum carriers from three yet unexplored malaria-mesoendemic areas with different transmission levels: two localities in northwestern Brazil (Ariquemes and Porto Velho) and a village in Madagascar (Ankazobe). A total of 89 DNA samples were analyzed by amplification of polymorphic domains from genes encoding merozoite surface antigens 1 and 2 (MSP-1, MSP-2) and thrombospondin-related anonymous protein (TRAP) and by hybridization with allelic-family-specific probes or random-fragment-length polymorphism (RFLP). In all three localities, extensive polymorphism was observed for each marker, but the MSP-2 central repeat was the most diverse one. Similar levels of genetic diversity, allelic frequency, and infection complexity were observed in the two Brazilian localities, although the isolates had been sampled at 2-year intervals, suggesting the stability of the infecting parasite populations presenting in these regions of the Brazilian Amazon. Unexpectedly, although the entomologic inoculation rate was at least 3 times lower in Ankazobe than in the Brazilian areas. Malagasi samples appeared more complex than the Brazilian ones. The implications of these data with regard to parasite population-dynamics studies are discussed.

Animals↗