Leishmaniases are vector-borne parasitic diseases with 0.9 – 1.4 million new human cases each year worldwide. In the vectorial part of the life-cycle, Leishmania development is confined to the digestive tract. During the first few days after blood feeding, natural barriers to Leishmania development include secreted proteolytic enzymes, the peritrophic matrix surrounding the ingested blood meal and sand fly immune reactions. As the blood digestion proceeds, parasites need to bind to the midgut epithelium to avoid being excreted with the blood remnant. This binding is strictly stage-dependent as it is a property of nectomonad and leptomonad forms only. While the attachment in specific vectors (P. papatasi, P. duboscqi and P. sergenti) involves lipophosphoglycan (LPG), this Leishmania molecule is not required for parasite attachment in other sand fly species experimentally permissive for various Leishmania. During late-stage infections, large numbers of parasites accumulate in the anterior midgut and produce filamentous proteophosphoglycan creating a gel-like plug physically obstructing the gut. The parasites attached to the stomodeal valve cause damage to the chitin lining and epithelial cells of the valve, interfering with its function and facilitating reflux of parasites from the midgut. Transformation to metacyclic stages highly infective for the vertebrate host is the other prerequisite for effective transmission. Here, we review the current state of knowledge of molecular interactions occurring in all these distinct phases of parasite colonization of the sand fly gut, highlighting recent discoveries in the field.

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Leishmania development in sand flies: parasite-vector interactions overview

The sexual phase of the malaria pathogen, Plasmodium falciparum, culminates in fertilization within the midgut of the mosquito and represents a crucial step in the completion of the parasite's life-cycle and transmission of the disease. Two decades ago, the first sexual stage-specific surface proteins were identified, among them Pfs230, Pfs48/45, and Pfs25, which were of scientific interest as candidates for the development of transmission blocking vaccines. A decade later, gene information gained from the sequencing of the P. falciparum genome led to the identification of numerous additional sexual-stage proteins with antigenic properties and novel enzymes that putatively possess regulatory functions during sexual-stage development. This review aims to summarize the sexual-stage proteins identified to date, to compare their stage specificities and expression patterns and to highlight novel regulative mechanisms of sexual differentiation. The prospective candidacy of select sexual-stage proteins as targets for transmission blocking strategies will be discussed.

Proteins of the malaria parasite sexual stages: expression, function and potential for transmission blocking strategies

Malaria transmission-blocking vaccines (TBVs) target the development of Plasmodium parasites within the mosquito, with the aim of preventing malaria transmission from one infected individual to another. Different vaccine platforms, mainly protein-in-adjuvant formulations delivering the leading candidate antigens, have been developed independently and have reported varied transmission-blocking activities (TBA). Here, recombinant chimpanzee adenovirus 63, ChAd63, and modified vaccinia virus Ankara, MVA, expressing AgAPN1, Pfs230-C, Pfs25, and Pfs48/45 were generated. Antibody responses primed individually against all antigens by ChAd63 immunization in BALB/c mice were boosted by the administration of MVA expressing the same antigen. These antibodies exhibited a hierarchy of inhibitory activity against the NF54 laboratory strain of P. falciparum in Anopheles stephensi mosquitoes using the standard membrane feeding assay (SMFA), with anti-Pfs230-C and anti-Pfs25 antibodies giving complete blockade. The observed rank order of inhibition was replicated against P. falciparum African field isolates in A. gambiae in direct membrane feeding assays (DMFA). TBA achieved was IgG concentration dependent. This study provides the first head-to-head comparative analysis of leading antigens using two different parasite sources in two different vector species, and can be used to guide selection of TBVs for future clinical development using the viral-vectored delivery platform.

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Comparative assessment of transmission-blocking vaccine candidates against Plasmodium falciparum.

The continued global burden of malaria can in part be attributed to a complex lifecycle, with both human hosts and mosquito vectors serving as transmission reservoirs. In preclinical models of vaccine-induced immunity, antibodies to parasite sexual-stage antigens, ingested in the mosquito blood meal, can inhibit parasite survival in the insect midgut as judged by ex vivo functional studies such as the membrane feeding assay. In an era of renewed political momentum for malaria elimination and eradication campaigns, such observations have fueled support for the development and implementation of so-called transmission-blocking vaccines. While leading candidates are being evaluated using a variety of promising vaccine platforms, the field is also beginning to capitalize on global '-omics' data for the rational genome-based selection and unbiased characterization of parasite and mosquito proteins to expand the candidate list. This review covers the progress and prospects of these recent developments.

Toward the development of effective transmission-blocking vaccines for malaria

Flipping the paradigm on malaria transmission-blocking vaccines

Anopheles gambiae is the major African vector of Plasmodium falciparum, the most deadly species of human malaria parasite and the most prevalent in Africa. Several strategies are being developed to limit the global impact of malaria via reducing transmission rates, among which are transmission-blocking vaccines (TBVs), which induce in the vertebrate host the production of antibodies that inhibit parasite development in the mosquito midgut. So far, the most promising components of a TBV are parasite-derived antigens, although targeting critical mosquito components might also successfully block development of the parasite in its vector. We previously identified A. gambiae genes whose expression was modified in P. falciparum-infected mosquitoes, including one midgut carboxypeptidase gene, cpbAg1. Here we show that P. falciparum up-regulates the expression of cpbAg1 and of a second midgut carboxypeptidase gene, cpbAg2, and that this up-regulation correlates with an increased carboxypeptidase B (CPB) activity at a time when parasites establish infection in the mosquito midgut. The addition of antibodies directed against CPBAg1 to a P. falciparum-containing blood meal inhibited CPB activity and blocked parasite development in the mosquito midgut. Furthermore, the development of the rodent parasite Plasmodium berghei was significantly reduced in mosquitoes fed on infected mice that had been immunized with recombinant CPBAg1. Lastly, mosquitoes fed on anti-CPBAg1 antibodies exhibited reduced reproductive capacity, a secondary effect of a CPB-based TBV that could likely contribute to reducing Plasmodium transmission. These results indicate that A. gambiae CPBs could constitute targets for a TBV that is based upon mosquito molecules.

https://iai.asm.org/content/iai/75/4/1635.full.pdf

A. Diop

Papers

Carboxypeptidases B of Anopheles gambiae as targets for a Plasmodium falciparum transmission-blocking vaccine.