scholarly journals Construction of a tetR-Integrated Salmonella enterica Serovar Typhi CVD908 Strain That Tightly Controls Expression of the Major Merozoite Surface Protein of Plasmodium falciparum for Applications in Human Vaccine Production

2002 ◽  
Vol 70 (4) ◽  
pp. 2029-2038 ◽  
Author(s):  
Feng Qian ◽  
Weiqing Pan
Keyword(s):  
Plasmodium Falciparum ◽  
Vaccine Strain ◽  
Salmonella Enterica ◽  
Expression Vector ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Inducible Expression ◽  
Luciferase Reporter ◽  
Foreign Gene

ABSTRACT Attenuated Salmonella strains are an attractive live vector for delivery of a foreign antigen to the human immune system. However, the problem with this vector lies with plasmid segregation and the low level of expression of the foreign gene in vivo when constitutive expression is employed, leading to a diminished immune response. We have established inducible expressions of foreign genes in the Salmonella enterica serovar Typhi CVD908 vaccine strain using the tetracycline response regulatory promoter. To set up this system, a tetracycline repressor (tetR) was integrated into a defined ΔaroC locus of the chromosome via suicide plasmid pJG12/tetR-neo. To remove the neo gene conferring kanamycin resistance from the locus, a cre expression vector under the control of the tetracycline response promoter was transformed into the clone; expression of the Cre recombinase excised the neo gene and generated the end strain CVD908-tetR. Expression of the luciferase reporter gene in this strain is dependent on the presence of tetracycline in the medium and can be regulated up to 4,773-fold. Moreover, the tightly controlled expression of major merozoite surface protein 1 (MSP1) and parts of Plasmodium falciparum was achieved, and the product yield was increased when the inducible expression system was employed. Inoculation of bacteria harboring plasmid pZE11/MSP142 in mice produced the protein in liver and spleen controlled by the inducer. The persistence of the plasmid-carrying bacteria in mice was determined. Peak colonization of both liver and spleen was detected on the third day postinoculation and was followed by a decline in growth curves. After 14 days postinfection, the majority of the bacteria (>90%) recovered from the liver and spleen of the mice retained the plasmid when expression was induced; this clearly indicated that stability of the expression vector in vivo was improved by inducible expression. Establishment of the regulatory system in the vaccine strain may broaden the range of its use by enhancing plasmid stability and expression levels in vivo. Moreover, the availability of the vaccine strain inducibly expressing the entire MSP1 provides possibilities for examining its immunogenicity, particularly the cellular response in animal models.

scholarly journals Immunoglobulin G Subclass-Specific Responses against Plasmodium falciparum Merozoite Antigens Are Associated with Control of Parasitemia and Protection from Symptomatic Illness

2009 ◽  
Vol 77 (3) ◽  
pp. 1165-1174 ◽  
Author(s):  
Danielle I. Stanisic ◽  
Jack S. Richards ◽  
Fiona J. McCallum ◽  
Pascal Michon ◽  
Christopher L. King ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Immunoglobulin G ◽  
Vaccine Development ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Igg Subclass ◽  
Apical Membrane Antigen 1 ◽  
Falciparum Merozoite ◽  
Merozoite Antigens

ABSTRACT Substantial evidence indicates that antibodies to Plasmodium falciparum merozoite antigens play a role in protection from malaria, although the precise targets and mechanisms mediating immunity remain unclear. Different malaria antigens induce distinct immunoglobulin G (IgG) subclass responses, but the importance of different responses in protective immunity from malaria is not known and the factors determining subclass responses in vivo are poorly understood. We examined IgG and IgG subclass responses to the merozoite antigens MSP1-19 (the 19-kDa C-terminal region of merozoite surface protein 1), MSP2 (merozoite surface protein 2), and AMA-1 (apical membrane antigen 1), including different polymorphic variants of these antigens, in a longitudinal cohort of children in Papua New Guinea. IgG1 and IgG3 were the predominant subclasses of antibodies to each antigen, and all antibody responses increased in association with age and exposure without evidence of increasing polarization toward one subclass. The profiles of IgG subclasses differed somewhat for different alleles of MSP2 but not for different variants of AMA-1. Individuals did not appear to have a propensity to make a specific subclass response irrespective of the antigen. Instead, data suggest that subclass responses to each antigen are generated independently among individuals and that antigen properties, rather than host factors, are the major determinants of IgG subclass responses. High levels of AMA-1-specific IgG3 and MSP1-19-specific IgG1 were strongly predictive of a reduced risk of symptomatic malaria and high-density P. falciparum infections. However, no antibody response was significantly associated with protection from parasitization per se. Our findings have major implications for understanding human immunity and for malaria vaccine development and evaluation.

scholarly journals Immunogenicity and In Vivo Efficacy of Recombinant Plasmodium falciparum Merozoite Surface Protein-1 in Aotus Monkeys

1995 ◽  
Vol 1 (3) ◽  
pp. 325-332 ◽  
Author(s):  
Sanjai Kumar ◽  
Anjali Yadava ◽  
David B. Keister ◽  
Jing Hui Tian ◽  
Michael Ohl ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
In Vivo Efficacy ◽  
Merozoite Surface Protein 1 ◽  
Falciparum Merozoite

scholarly journals Biochemical and Immunological Characterization of Bacterially Expressed and Refolded Plasmodium falciparum 42-Kilodalton C-Terminal Merozoite Surface Protein 1

2003 ◽  
Vol 71 (12) ◽  
pp. 6766-6774 ◽  
Author(s):  
Sanjay Singh ◽  
Michael C. Kennedy ◽  
Carole A. Long ◽  
Allan J. Saul ◽  
Louis H. Miller ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Native Form ◽  
Immunological Characterization ◽  
E Coli ◽  
Merozoite Surface Protein 1 ◽  
Vaccine Trials

ABSTRACT Protection against Plasmodium falciparum can be induced by vaccination in animal models with merozoite surface protein 1 (MSP1), which makes this protein an attractive vaccine candidate for malaria. In an attempt to produce a product that is easily scaleable and inexpensive, we expressed the C-terminal 42 kDa of MSP1 (MSP142) in Escherichia coli, refolded the protein to its native form from insoluble inclusion bodies, and tested its ability to elicit antibodies with in vitro and in vivo activities. Biochemical, biophysical, and immunological characterization confirmed that refolded E. coli MSP142 was homogeneous and highly immunogenic. In a formulation suitable for human use, rabbit antibodies were raised against refolded E. coli MSP142 and tested in vitro in a P. falciparum growth invasion assay. The antibodies inhibited the growth of parasites expressing either homologous or heterologous forms of P. falciparum MSP142. However, the inhibitory activity was primarily a consequence of antibodies directed against the C- terminal 19 kDa of MSP1 (MSP119). Vaccination of nonhuman primates with E. coli MSP142 in Freund's adjuvant protected six of seven Aotus monkeys from virulent infection with P. falciparum. The protection correlated with antibody-dependent mechanisms. Thus, this new construct, E. coli MSP142, is a viable candidate for human vaccine trials.

Disruption of Plasmodium berghei merozoite surface protein 7 gene modulates parasite growth in vivo

Blood ◽  
2005 ◽  
Vol 105 (1) ◽  
pp. 394-396 ◽  
Author(s):  
Rita Tewari ◽  
Solabomi A. Ogun ◽  
Ruwani S. Gunaratne ◽  
Andrea Crisanti ◽  
Anthony A. Holder
Keyword(s):  
Plasmodium Falciparum ◽  
Gene Family ◽  
Plasmodium Berghei ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Plasmodium Yoelii ◽  
Parasite Growth ◽  
Surface Proteins ◽  
Rate Of Increase

Abstract Merozoite invasion of red blood cells is crucial to the development of the parasite that causes malaria. Merozoite surface proteins (MSPs) mediate the first interaction between parasite and erythrocyte. In Plasmodium falciparum, they include a complex of products from at least 3 genes (msp1, msp6, and msp7), one of which, msp7, is part of a gene family containing 3 and 6 adjacent members in Plasmodium yoelii and Plasmodium falciparum, respectively. We have identified and disrupted msp7 in the Plasmodium berghei gene family. The protein is expressed in schizonts and colocalizes with MSP1. The synthesis and processing of MSP1 was unaffected in the parasite with the disrupted gene (MSP7ko). Disruption of msp7 was not lethal but affected blood-stage parasite growth. MSP7ko parasites initially grew more slowly than wild-type parasites. However, when reticulocytes were prevalent, the rate of increase in parasitemia was similar, suggesting that MSP7ko parasites prefer to invade and grow within reticulocytes. (Blood. 2005;105:394-396)

scholarly journals Vaccination of Monkeys with Recombinant Plasmodium falciparum Apical Membrane Antigen 1 Confers Protection against Blood-Stage Malaria

2002 ◽  
Vol 70 (12) ◽  
pp. 6961-6967 ◽  
Author(s):  
Anthony W. Stowers ◽  
Michael C. Kennedy ◽  
Brian P. Keegan ◽  
Allan Saul ◽  
Carole A. Long ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Malaria Vaccine ◽  
Vaccine Development ◽  
Apical Membrane ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Lethal Challenge ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen

ABSTRACT A major challenge facing malaria vaccine development programs is identifying efficacious combinations of antigens. To date, merozoite surface protein 1 (MSP1) is regarded as the leading asexual vaccine candidate. Apical membrane antigen 1 (AMA1) has been identified as another leading candidate for an asexual malaria vaccine, but without any direct in vivo evidence that a recombinant form of Plasmodium falciparum AMA1 would have efficacy. We evaluated the efficacy of a form of P. falciparum AMA1, produced in Pichia pastoris, by vaccinating Aotus vociferans monkeys and then challenging them with P. falciparum parasites. Significant protection from this otherwise lethal challenge with P. falciparum was observed. Five of six animals had delayed patency; two of these remained subpatent for the course of the infection, and two controlled parasite growth at <0.75% of red blood cells parasitized. The protection induced by AMA1 was superior to that obtained with a form of MSP1 used in the same trial. The protection induced by a combination vaccine of AMA1 and MSP1 was not superior to the protection obtained with AMA1 alone, although the immunity generated appeared to operate against both vaccine components.

scholarly journals Efficacy of Two Alternate Vaccines Based on Plasmodium falciparum Merozoite Surface Protein 1 in anAotus Challenge Trial

2001 ◽  
Vol 69 (3) ◽  
pp. 1536-1546 ◽  
Author(s):  
Anthony W. Stowers ◽  
Vittoria Cioce ◽  
Richard L. Shimp ◽  
Mark Lawson ◽  
George Hui ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Vaccine Antigen ◽  
Merozoite Surface Protein 1 ◽  
Adjuvant Vaccine ◽  
New Formulation ◽  
Antibody Levels

ABSTRACT In an attempt to produce a more defined, clinical-grade version of a vaccine based on Plasmodium falciparum merozoite surface protein 1 (MSP1), we evaluated the efficacy of two recombinant forms of MSP1 in an Aotus nancymai challenge model system. One recombinant vaccine, bvMSP142, based on the 42-kDa C-terminal portion of MSP1, was expressed as a secreted protein in baculovirus-infected insect cells. A highly pure baculovirus product could be reproducibly expressed and purified at yields in excess of 8 mg of pure protein per liter of culture. This protein, when tested for efficacy in the Aotus challenge model, gave significant protection, with only one of seven monkeys requiring treatment for uncontrolled parasitemia after challenge with P. falciparum. The second recombinant protein, P30P2MSP119, has been used in previous studies and is based on the smaller, C-terminal 19-kDa portion of MSP1 expressed inSaccharomyces cerevisiae. Substantial changes were made in its production process to optimize expression. The optimum form of this vaccine antigen (as judged by in vitro and in vivo indicators) was then evaluated, along with bvMSP142, for efficacy in theA. nancymai system. The new formulation of P30P3MSP119 performed significantly worse than bvMSP142 and appeared to be less efficacious than we have found in the past, with four of seven monkeys in the vaccinated group requiring treatment for uncontrolled parasitemia. With both antigens, protection was seen only when high antibody levels were obtained by formulation of the vaccines in Freund's adjuvant. Vaccine formulation in an alternate adjuvant, MF59, resulted in significantly lower antibody titers and no protection.

scholarly journals Quantification of Plasmodium falciparum malaria from complex infections in the Peruvian Amazon using quantitative PCR of the merozoite surface protein 1, block 2 (PfMSP1-B2): in vitro dynamics reveal density-dependent interactions

Parasitology ◽  
2012 ◽  
Vol 139 (6) ◽  
pp. 701-708 ◽  
Author(s):  
THOMAS M. ZERVOS ◽  
JEAN N. HERNANDEZ ◽  
PATRICK L. SUTTON ◽  
ORALEE H. BRANCH
Keyword(s):  
Plasmodium Falciparum ◽  
Quantitative Pcr ◽  
Surface Protein ◽  
Plasmodium Falciparum Malaria ◽  
Merozoite Surface Protein ◽  
Peruvian Amazon ◽  
Field Isolates ◽  
Merozoite Surface Protein 1

SUMMARYThe majority of Plasmodium falciparum field isolates are defined as complex infections because they contain multiple genetically distinct clones. Studying interactions between clones in complex infections in vivo and in vitro could elucidate important phenomena in malaria infection, transmission and treatment. Using quantitative PCR (qPCR) of the P. falciparum merozoite surface protein 1, block 2 (PfMSP1-B2), we provide a sensitive and efficient genotyping method. This is important for epidemiological studies because it makes it possible to study genotype-specific growth dynamics. We compared 3 PfMSP1-B2 genotyping methods by analysing 79 field isolates from the Peruvian Amazon. In vivo observations from other studies using these techniques led to the hypothesis that clones within complex infections interact. By co-culturing clones with different PfMSP1-B2 genotypes, and measuring parasitaemia using qPCR, we found that suppression of clonal expansion was a factor of the collective density of all clones present in a culture. PfMSP1-B2 qPCR enabled us to find in vitro evidence for parasite-parasite interactions and could facilitate future investigations of growth trends in naturally occurring complex infections.

scholarly journals A New Rodent Model to Assess Blood Stage Immunity to the Plasmodium falciparum Antigen Merozoite Surface Protein 119 Reveals a Protective Role for Invasion Inhibitory Antibodies

2003 ◽  
Vol 198 (6) ◽  
pp. 869-875 ◽  
Author(s):  
Tania F. de Koning-Ward ◽  
Rebecca A. O'Donnell ◽  
Damien R. Drew ◽  
Russell Thomson ◽  
Terence P. Speed ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Malaria Parasite ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Protective Role ◽  
Blood Stage ◽  
Parasite Line ◽  
Parasite Plasmodium ◽  
Inhibitory Antibodies

Antibodies capable of inhibiting the invasion of Plasmodium merozoites into erythrocytes are present in individuals that are clinically immune to the malaria parasite. Those targeting the 19-kD COOH-terminal domain of the major merozoite surface protein (MSP)-119 are a major component of this inhibitory activity. However, it has been difficult to assess the overall relevance of such antibodies to antiparasite immunity. Here we use an allelic replacement approach to generate a rodent malaria parasite (Plasmodium berghei) that expresses a human malaria (Plasmodium falciparum) form of MSP-119. We show that mice made semi-immune to this parasite line generate high levels of merozoite inhibitory antibodies that are specific for P. falciparum MSP-119. Importantly, protection from homologous blood stage challenge in these mice correlated with levels of P. falciparum MSP-119–specific inhibitory antibodies, but not with titres of total MSP-119–specific immunoglobulins. We conclude that merozoite inhibitory antibodies generated in response to infection can play a significant role in suppressing parasitemia in vivo. This study provides a strong impetus for the development of blood stage vaccines designed to generate invasion inhibitory antibodies and offers a new animal model to trial P. falciparum MSP-119 vaccines.

scholarly journals Allelic diversity of merozoite surface protein genes (msp1 and msp2) and clinical manifestations of Plasmodium falciparum malaria cases in Aceh, Indonesia

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Kurnia Fitri Jamil ◽  
Nandha Rizki Pratama ◽  
Sylvia Sance Marantina ◽  
Harapan Harapan ◽  
Muhammad Riza Kurniawan ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Liver Function ◽  
Severe Malaria ◽  
Clinical Manifestation ◽  
Allelic Diversity ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Malaria Control Programme ◽  
Co Morbidity ◽  
Aceh Province

Abstract Background The malaria control programme in Indonesia has successfully brought down malaria incidence in many parts in Indonesia, including Aceh Province. Clinical manifestation of reported malaria cases in Aceh varied widely from asymptomatic, mild uncomplicated to severe and fatal complications. The present study aims to explore the allelic diversity of merozoite surface protein 1 gene (msp1) and msp2 among the Plasmodium falciparum isolates in Aceh Province and to determine their potential correlation with the severity of malaria clinical manifestation. Methods Screening of over 500 malaria cases admitted to the hospitals in 11 districts hospital within Aceh Province during 2013–2015, identified 90 cases of P. falciparum mono-infection without any co-morbidity. The subjects were clinically phenotyped and parasite DNA was extracted and polymerase chain reaction (PCR) amplified for the msp1 and msp2 allelic subfamilies. Results Analysis of clinical manifestation revealed that fever-chill is the most frequent symptom. Based on WHO criteria showed 19 cases were classified as severe and 71 as mild malaria. Analysis of msp1 gene revealed the presence of K1 allele subfamily in 34 subjects, MAD20 in 42 subjects, RO33 in 1 subject, and mixed allelic of K1 + MAD20 in 5 subjects, K1 + RO33 in 4 subjects, and MAD20 + RO33 in 4 subjects. Analysis of msp2 gene revealed 34 subjects carried the FC27 allelic subfamily, 37 subjects carried the 3D7 and 19 subjects carried the mixed FC27 + 3D7. Analysis of multiplicity of infection revealed that msp1 alleles is slightly higher than msp2 with the mean of MOI were 2.69 and 2.27, respectively. Statistical analysis to determine the association between each clinical manifestation and msp1 and msp2 alleles revealed that liver function abnormal value was associated with the msp2 mixed alleles (odds ratio (OR):0.13; 95%CI: 0.03–0.53). Mixed msp1 of K1 + RO33 was associated with severe malaria (OR: 28.50; 95%CI: 1.59–1532.30). Conclusion This study found a strong association between severe malaria in Aceh with subjects carrying the msp1 mixed alleles of K1 and RO33. The liver function abnormal value associated with the msp2 mixed allelic subfamilies. Further study in different geographic areas is recommended.

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