Cellular responses to modified Plasmodium falciparum MSP119 antigens in individuals previously exposed to natural malaria infection

<p>Abstract</p> <p>Background</p> <p>MSP1 processing-inhibitory antibodies bind to epitopes on the 19 kDa C-terminal region of the <it>Plasmodium falciparum </it>merozoite surface protein 1 (MSP1₁₉), inhibiting erythrocyte invasion. Blocking antibodies also bind to this antigen but prevent inhibitory antibodies binding, allowing invasion to proceed. Recombinant MSP1₁₉had been modified previously to allow inhibitory but not blocking antibodies to continue to bind. Immunization with these modified proteins, therefore, has the potential to induce more effective protective antibodies. However, it was unclear whether the modification of MSP1₁₉would affect critical T-cell responses to epitopes in this antigen.</p> <p>Methods</p> <p>The cellular responses to wild-type MSP1₁₉and a panel of modified MSP1₁₉antigens were measured using an <it>in-vitro </it>assay for two groups of individuals: the first were malaria-naïve and the second had been naturally exposed to <it>Plasmodium falciparum </it>infection. The cellular responses to the modified proteins were examined using cells from malaria-exposed infants and adults.</p> <p>Results</p> <p>Interestingly, stimulation indices (SI) for responses induced by some of the modified proteins were at least two-fold higher than those elicited by the wild-type MSP1₁₉. A protein with four amino acid substitutions (Glu27→Tyr, Leu31→Arg, Tyr34→Ser and Glu43→Leu) had the highest stimulation index (SI up to 360) and induced large responses in 64% of the samples that had significant cellular responses to the modified proteins.</p> <p>Conclusion</p> <p>This study suggests that specific MSP1₁₉variants that have been engineered to improve their antigenicity for inhibitory antibodies, retain T-cell epitopes and the ability to induce cellular responses. These proteins are candidates for the development of MSP1-based malaria vaccines.</p

Formation of the Food Vacuole in Plasmodium falciparum: A Potential Role for the 19 kDa Fragment of Merozoite Surface Protein 1 (MSP119)

Plasmodium falciparum Merozoite Surface Protein 1 (MSP1) is synthesized during schizogony as a 195-kDa precursor that is processed into four fragments on the parasite surface. Following a second proteolytic cleavage during merozoite invasion of the red blood cell, most of the protein is shed from the surface except for the C-terminal 19-kDa fragment (MSP119), which is still attached to the merozoite via its GPI-anchor. We have examined the fate of MSP119 during the parasite's subsequent intracellular development using immunochemical analysis of metabolically labeled MSP119, fluorescence imaging, and immuno-electronmicroscopy. Our data show that MSP119 remains intact and persists to the end of the intracellular cycle. This protein is the first marker for the biogenesis of the food vacuole; it is rapidly endocytosed into small vacuoles in the ring stage, which coalesce to form the single food vacuole containing hemozoin, and persists into the discarded residual body. The food vacuole is marked by the presence of both MSP119 and the chloroquine resistance transporter (CRT) as components of the vacuolar membrane. Newly synthesized MSP1 is excluded from the vacuole. This behavior indicates that MSP119 does not simply follow a classical lysosome-like clearance pathway, instead, it may play a significant role in the biogenesis and function of the food vacuole throughout the intra-erythrocytic phase

Genome-wide profiling of humoral immunity and pathogen genes under selection identifies immune evasion tactics of Chlamydia trachomatis during ocular infection

The frequency and duration of Chlamydia trachomatis (Ct) ocular infections decrease with age, suggesting development of partial immunity. However, there is a lack of clear correlates of immunity to Ct infection in humans. We screened sera from a cohort of Gambian children followed for six-months against a Ct-proteome microarray. At genome sequence level, we detected signatures of selection from a population of ocular Ct isolates from Guinea-Bissau. Together these approaches allowed us to highlight the focus of humoral responses and hypothesise new modes of pathogen immune evasion. Children who were susceptible to frequent and/or prolonged Ct infection had a less focussed antibody response, including preferential recognition of forty-two antigens. There was evidence of positive and purifying selection across the genome, but little balancing selection. In contrast, most antigens that were associated with susceptibility were under neutral selection. These data suggest an evasion strategy in which Ct presents a large panel of irrelevant antigens to the immune system to block or misdirect protective responses. Development of a focused immune response, possibly induced through vaccination, may be an effective strategy to promote protection to Ct infection

Effects of Mowing on Methane Uptake in a Semiarid Grassland in Northern China

Background: Mowing is a widely adopted management practice for the semiarid steppe in China and affects CH4 exchange. However, the magnitude and the underlying mechanisms for CH 4 uptake in response to mowing remain uncertain. Methodology/Principal Findings: In two consecutive growing seasons, we measured the effect of mowing on CH 4 uptake in a steppe community. Vegetation was mowed to 2 cm (M2), 5 cm (M5), 10 cm (M10), 15 cm (M15) above soil surface, respectively, and control was set as non-mowing (NM). Compared with control, CH4 uptake was substantially enhanced at almost all the mowing treatments except for M15 plots of 2009. CH4 uptake was significantly correlated with soil microbial biomass carbon, microbial biomass nitrogen, and soil moisture. Mowing affects CH 4 uptake primarily through its effect on some biotic factors, such as net primary productivity, soil microbial C\N supply and soil microbial activities, while soil temperature and moisture were less important. Conclusions/Significance: This study found that mowing affects the fluxes of CH4 in the semiarid temperate steppe of north China

A principal target of human immunity to malaria identified by molecular population genetic and immunological analyses

New strategies are required to identify the most important targets of protective immunity in complex eukaryotic pathogens. Natural selection maintains allelic variation in some antigens of the malaria parasite Plasmodium falciparum (1–3). Analysis of allele frequency distributions could identify the loci under most intense selection (4–7). The merozoite surface protein 1 (Msp1) is the most-abundant surface component on the erythrocyte-invading stage of P. falciparum (8–10). Immunization with whole Msp1 has protected monkeys completely against homologous (11) and partially against non-homologous (12) parasite strains. The singlecopy msp1 gene, of about 5 kilobases, has highly divergent alleles (13) with stable frequencies in endemic populations (14,15). To identify the region of msp1 under strongest selection to maintain alleles within populations, we studied multiple intragenic sequence loci in populations in different regions of Africa and Southeast Asia. On both continents, the locus with the lowest inter-population variance in allele frequencies was block 2, indicating selection in this part of the gene. To test the hypothesis of immune selection, we undertook a large prospective longitudinal cohort study. This demonstrated that serum IgG antibodies against each of the two most frequent allelic types of block 2 of the protein were strongly associated with protection from P. falciparum malaria

New insights for native production of MSP1(19), the disulfide-rich C-terminal fragment from Plasmodium falciparum merozoite surface protein 1.

Malaria represents a major public health problem and an important cause of mortality and morbidity. The malaria parasites are becoming resistant to drugs used to treat the disease and still no efficient vaccine has been developed. One promising vaccine candidate is the merozoite surface protein 1 (MSP1), which has been extensively investigated as a vaccine target. The surface protein MSP1 plays an essential role in the erythrocyte invasion process and is an accessible target for the immune system. Antibodies to the carboxy-terminal region of the protein, named MSP119, can inhibit erythrocyte invasion and parasite growth. In order to develop an effective MSP119- based vaccine against malaria, production of an antigen that is recognized by protective antibodies is mandatory. To this aim, we propose a method to produce the disulfide-rich MSP119 in its native conformation based on its in vitro oxidative refolding. The native conformation of the renatured MSP119 is carefully established by immunochemical reactivity experiments, circular dichroism and NMR. MSP119 can successfully be refolded in vitro as an isolated protein or as a fusion with the maltose binding protein. The possibility to properly fold MSP119in vitro paves the way to new approaches for high titer production of native MSP119 using Escherichia coli as a host