Retrospective study of toxoplasmosis prevalence in pregnant women in Benin and its relation with malaria

Background Globally distributed with variable prevalence depending on geography, toxoplasmosis is a zoonosis caused by an obligate intracellular protozoan parasite, Toxoplasma gondii. This disease is usually benign but poses a risk for immunocompromised people and for newborns of mothers with a primary infection during pregnancy because of the risk of congenital toxoplasmosis (CT). CT can cause severe damage to fetuses-newborns. To our knowledge, no study has been conducted in sub-Saharan Africa on toxoplasmosis seroprevalence, seroconversion and CT in a large longitudinal cohort and furthermore, no observation has been made of potential relationships with malaria. Methods We performed a retrospective toxoplasmosis serological study using available samples from a large cohort of 1,037 pregnant women who were enrolled in a malaria follow-up during the 2008–2010 period in a rural area in Benin. We also used some existing data to investigate potential relationships between the maternal toxoplasmosis serological status and recorded malaria infections. Results Toxoplasmosis seroprevalence, seroconversion and CT rates were 52.6%, 3.4% and 0.2%, respectively, reflecting the population situation of toxoplasmosis, without targeted medical intervention. The education level influences the toxoplasmosis serological status of women, with women with little or no formal education have greater immunity than others. Surprisingly, toxoplasmosis seropositive pregnant women tended to present lower malaria infection during pregnancy (number) or at delivery (presence) and to have lower IgG levels to Plasmodium falciparum Apical Membrane Antigen 1, compared to toxoplasmosis seronegative women. Conclusions The high toxoplasmosis seroprevalence indicates that prevention against this parasite remains important to deploy and must be accessible and understandable to and for all individuals (educated and non-educated). A potential protective role against malaria conferred by a preexisting toxoplasmosis infection needs to be explored more precisely to examine the environmental, parasitic and/or immune aspects.

Genetic Diversity and Selection of Plasmodium vivax Apical Membrane Antigen-1 in China–Myanmar Border of Yunnan Province, China, 2009–2016

Plasmodium vivax apical membrane antigen-1 (PvAMA-1) is an important vaccine candidate for vivax malaria. However, antigenic variation within PvAMA-1 is a major obstacle to the design of a global protective malaria vaccine. In this study, we analyzed the genetic polymorphism and selection of the PvAMA-1 gene from 152 P. vivax isolates from imported cases to China, collected in the China–Myanmar border (CMB) area in Yunnan Province (YP) during 2009–2011 (n = 71) and 2014–2016 (n = 81), in comparison with PvAMA-1 gene information from Myanmar (n = 73), collected from public data. The overall nucleotide diversity of the PvAMA-1 gene from the 152 YP isolates was 0.007 with 76 haplotypes identified (Hd = 0.958). Results from the population structure suggested three groups among the YP and Myanmar isolates with optimized clusters value of K = 7. In addition, YP (2014–2016) isolates generally lacked some K components that were commonly found in YP (2009–2011) and Myanmar. Meanwhile, PvAMA-1 domain I is found to be the dominant target of positive diversifying selection and most mutation loci were found in this domain. The mutation frequencies of D107N/A, R112K/T, K120R, E145A, E277K, and R438H in PvAMA-1 were more than 70% in the YP isolates. In conclusion, high genetic diversity and positive selection were found in the PvAMA-1 gene from YP isolates, which are significant findings for the design and development of PvAMA-1-based malaria vaccine.

The AMA1-RON complex drives Plasmodium sporozoite invasion in the mosquito and mammalian hosts

Plasmodium sporozoites that are transmitted by blood-feeding female Anopheles mosquitoes invade hepatocytes for an initial round of intracellular replication, leading to the release of merozoites that invade and multiply within red blood cells. Sporozoites and merozoites share a number of proteins that are expressed by both stages, including the Apical Membrane Antigen 1 (AMA1) and the Rhoptry Neck Proteins (RONs). Although AMA1 and RONs are essential for merozoite invasion of erythrocytes during asexual blood stage replication of the parasite, their function in sporozoites is still unclear. Here we show that AMA1 interacts with RONs in mature sporozoites. By using DiCre-mediated conditional gene deletion in P. berghei, we demonstrate that loss of AMA1, RON2 or RON4 in sporozoites impairs colonization of the mosquito salivary glands and invasion of mammalian hepatocytes, without affecting transcellular parasite migration. Our data establish that AMA1 and RONs facilitate host cell invasion across Plasmodium invasive stages, and suggest that sporozoites use the AMA1-RON complex to safely enter the mosquito salivary glands without causing cell damage, to ensure successful parasite transmission. These results open up the possibility of targeting the AMA1-RON complex for transmission-blocking antimalarial strategies.

Genetic Polymorphism and Natural Selection of Apical Membrane Antigen-1 in Plasmodium falciparum Isolates from Vietnam

Apical membrane antigen-1 of Plasmodium falciparum (PfAMA-1) is a leading malaria vaccine candidate antigen. However, the genetic diversity of pfama-1 and associated antigenic variation in global P. falciparum field isolates are major hurdles to the design of an efficacious vaccine formulated with this antigen. Here, we analyzed the genetic structure and the natural selection of pfama-1 in the P. falciparum population of Vietnam. A total of 37 distinct haplotypes were found in 131 P. falciparum Vietnamese isolates. Most amino acid changes detected in Vietnamese pfama-1 were localized in the ectodomain, domains I, II, and III. Overall patterns of major amino acid changes in Vietnamese pfama-1 were similar to those of global pfama-1, but the frequencies of the amino acid changes slightly differed by country. Novel amino acid changes were also identified in Vietnamese pfama-1. Vietnamese pfama-1 revealed relatively lower genetic diversity than currently analyzed pfama-1 in other geographical regions, and suggested a distinct genetic differentiation pattern. Evidence for natural selection was detected in Vietnamese pfama-1, but it showed purifying selection unlike the global pfama-1 analyzed so far. Recombination events were also found in Vietnamese pfama-1. Major amino acid changes that were commonly identified in global pfama-1 were mainly localized to predicted B-cell epitopes, RBC-binding sites, and IUR regions. These results provide important information for understanding the genetic nature of the Vietnamese pfama-1 population, and have significant implications for the design of a vaccine based on PfAMA-1.

The New Human Babesia sp. FR1 Is a European Member of the Babesia sp. MO1 Clade

In Europe, Babesia divergens is responsible for most of the severe cases of human babesiosis. In the present study, we describe a case of babesiosis in a splenectomized patient in France and report a detailed molecular characterization of the etiological agent, named Babesia sp. FR1, as well as of closely related Babesia divergens, Babesia capreoli and Babesia sp. MO1-like parasites. The analysis of the conserved 18S rRNA gene was supplemented with the analysis of more discriminant markers involved in the red blood cell invasion process: rap-1a (rhoptry-associated-protein 1) and ama-1 (apical-membrane-antigen 1). The rap-1a and ama-1 phylogenetic analyses were congruent, placing Babesia sp. FR1, the new European etiological agent, in the American cluster of Babesia sp. MO1-like parasites. Based on two additional markers, our analysis confirms the clear separation of B. divergens and B. capreoli. Babesia sp. MO1-like parasites should also be considered as a separate species, with the rabbit as its natural host, differing from those of B. divergens (cattle) and B. capreoli (roe deer). The natural host of Babesia sp. FR1 remains to be discovered.

A Chimeric Peptide Inhibits Red Blood Cell Invasion by Plasmodium falciparum with Hundredfold Increased Efficacy

Inhibition of tight junction formation between two malaria parasite proteins, apical membrane antigen 1 and rhoptry neck protein 2, crucial for red blood cell invasion, prevents the disease progression. In this work, we have utilized a unique approach to design a chimeric peptide, prepared by fusion of the best features of two peptide inhibitors, that has displayed parasite growth inhibition, in-vitro, with nanomolar IC50, which is hundredfold better than any of its parent peptides. Further, to gain structural insights, we computationally modeled the hybrid peptide on its receptor.

Quantification of the Dynamics of Antibody Response to Malaria to Inform Sero-Surveillance in Pregnant Women

Background: Malaria remains a major public health threat and in low malaria transmission areas new tools are needed to detect infections for prompt treatment and to progress elimination efforts. Pregnant women are particularly vulnerable to malaria infections and access routine antenatal care, presenting a unique sentinel population to apply novel sero-surveillance tools to measure malaria transmission. The aim of this study was to quantify the dynamic antibody responses to multiple antigens during pregnancy to identify a single or multiple antibody response of exposure to malaria in pregnancy. Methods: Antibody-mediated immunity responses to six parasite antigens (five commonly studied merozoite antigens and the variant surface antigen 2-chondroitin sulphate A (VAR2CSA), a pregnancy-specific erythrocytic antigen) were measured over the gestation period until delivery (median of 7 measurements/woman) in 250 pregnant women who attended antenatal clinics located at the Thai-Myanmar border. A multivariate mixture linear mixed model was used to cluster the pregnant women into groups that have similar longitudinal antibody responses to all six antigens over the gestational period using a Bayesian approach. The variable-specific entropy was calculated to identify the antibody responses that have the highest influence on the classification of the women into clusters, and subsequent agreement with grouping of women based on exposure to malaria during pregnancy. Results: Of the 250 pregnant women, 135 had a Plasmodium infection detected by light microscopy during pregnancy, defined as cases. The antibody responses to all six antigens accurately identified the women who did not have a malaria infection detected during pregnancy (93%, 107/115 controls). Antibody responses to P. falciparum merozoite surface protein 3 (PfMSP3) and P. vivax apical membrane antigen 1 (PvAMA1) were the least dynamic. Antibody responses to the antigens P. falciparum apical membrane antigen 1 (PfAMA1) and PfVAR2CSA were able to identify the majority of the cases more accurately (63%, 85/135). Conclusion: These findings suggest that the combination of antibodies, PfAMA1 and PfVAR2CSA, may be useful for sero-surveillance of malaria infections in pregnant women, particularly in low malaria transmission settings, leading to the early detection and treatment of malaria infections in pregnant women.

Structural and immunological characterization of an epitope within the PAN motif of ectodomain I in Babesia bovis apical membrane antigen 1 for vaccine development

Background Bovine babesiosis caused by Babesia bovis (B. bovis) has had a significant effect on the mobility and mortality rates of the cattle industry worldwide. Live-attenuated vaccines are currently being used in many endemic countries, but their wide use has been limited for a number of reasons. Although recombinant vaccines have been proposed as an alternative to live vaccines, such vaccines are not commercially available to date. Apical membrane antigen-1 (AMA-1) is one of the leading candidates in the development of a vaccine against diseases caused by apicomplexan parasite species. In Plasmodium falciparum (P. falciparum) AMA-1 (PfAMA-1), several antibodies against epitopes in the plasminogen, apple, and nematode (PAN) motif of PfAMA-1 domain I significantly inhibited parasite growth. Therefore, the purpose of this study was to predict an epitope from the PAN motif of domain I in the B. bovis AMA-1 (BbAMA-1) using a combination of linear and conformational B-cell epitope prediction software. The selected epitope was then bioinformatically analyzed, synthesized as a peptide (sBbAMA-1), and then used to immunize a rabbit. Subsequently, in vitro growth- and the invasion-inhibitory effects of the rabbit antiserum were immunologically characterized. Results Our results demonstrated that the predicted BbAMA-1 epitope was located on the surface-exposed α-helix of the PAN motif in domain I at the apex area between residues 181 and 230 with six polymorphic sites. Subsequently, sBbAMA-1 elicited antibodies capable of recognizing the native BbAMA-1 in immunoassays. Furthermore, anti-serum against sBbAMA-1 was immunologically evaluated for its growth- and invasion-inhibitory effects on B. bovis merozoites in vitro. Our results demonstrated that the rabbit anti-sBbAMA-1 serum at a dilution of 1:5 significantly inhibited (p < 0.05) the growth of B. bovis merozoites by approximately 50–70% on days 3 and 4 of cultivation, along with the invasion of merozoites by approximately 60% within 4 h of incubation when compared to the control groups. Conclusion Our results indicate that the epitope predicted from the PAN motif of BbAMA-1 domain I is neutralization-sensitive and may serve as a target antigen for vaccine development against bovine babesiosis caused by B. bovis.