Biology subjects

Alves e Silva, T. L.

Publications and source records attributed to Alves e Silva, T. L..

5 recordsLinked to original sources

bioRxiv2025

Unlocking new understanding of Plasmodium sporozoite biology with expansion microscopy

Transmission of malaria relies on the formation of sporozoites in the mosquito midgut and their subsequent invasion of the salivary gland. Despite their importance, our understanding of the cell biology of sporozoite formation and salivary gland invasion is limited. Here, we apply a technique called Mosquito Tissue Ultrastructure Expansion Microscopy (MoTissU-ExM), which physically expands infected mosquito tissues while preserving both host and parasite ultrastructure. Using MoTissU-ExM we are able to observe a range of parasite structures and organelles including features previously seen only by electron microscopy as well as structure not observed before. We leverage MoTissU-ExM to investigate a number of cell biology events during sporozoite formation and salivary gland invasion. In particular we focus on the rhoptries, a secretory organelle important for host cell invasion. We establish a timeline for sporozoite rhoptry biogenesis, show that two rhoptries are used up during salivary gland invasion, and provide the first evidence that rhoptry pairs are specialized for different invasion events. Building on this new knowledge, we characterize the rhoptry protein RON11 and identify it as the first protein involved in sporozoite rhoptry biogenesis. Disruption of RON11 led to the production of sporozoites that specifically fail to invade the salivary gland epithelial cell, thereby blocking transmission of these parasites.

microbiology↗

bioRxiv2025

High-Resolution Proteomics Unveils Salivary Gland Disruption and Saliva-Hemolymph Protein Exchange in Plasmodium-Infected Mosquitoes

Plasmodium sporozoites, the stage that initiates a malaria infection, must invade the mosquito salivary glands (SGs) before transmitting to a vertebrate host. However, the effects of sporozoite invasion on salivary gland physiology and saliva composition remain largely unexplored. We examined the impact of Plasmodium infection on Anopheles gambiae salivary glands using high-resolution proteomics, gene expression, and morphological analysis. The data revealed differential expression of various proteins, including the enrichment of humoral proteins in infected salivary glands originating from the hemolymph. These proteins diffused into the SGs due to structural damage caused by the sporozoites during invasion. Conversely, saliva proteins diffused out into the circulation of infected mosquitoes. Moreover, infection altered saliva protein composition, as shown by proteomes from saliva collected from mosquitoes infected by P. berghei or P. falciparum, revealing a significant reduction of immune proteins compared to uninfected mosquitoes. This reduction is likely due to the association of these proteins with the surface of sporozoites within the mosquito salivary secretory cavities. The saliva protein profiles from mosquitoes infected with both Plasmodium species were remarkably similar, suggesting a conserved interaction between sporozoites and salivary glands. Our results provide a foundation for understanding the molecular interactions between Plasmodium sporozoites and mosquito salivary glands.

microbiology↗

bioRxiv2024

Mosquito Tissue Ultrastructure-Expansion Microscopy (MoTissU-ExM) enables ultrastructural and anatomical analysis of malaria parasites and their mosquito.

Study of malaria parasite cell biology is challenged by their small size, which can make visualisation of individual organelles difficult or impossible using conventional light microscopy. In recent years, the [fi]eld has attempted to overcome this challenge through the application of ultrastructure expansion microscopy (U-ExM), which physically expands a biological sample approximately 4.5-fold. To date, U-ExM has mostly been used to visualise blood-stage parasites and used exclusively on parasites in vitro. Here we develop Mosquito Tissue U-ExM (MoTissU-ExM), a method for preparing dissected mosquito salivary glands and midguts by U-ExM. MoTissU-ExM preserves both host and parasite ultrastructure, enabling visualisation of oocysts and sporozoites in situ. We validate that MoTissU-ExM samples expand as expected, provide a direct comparison of the same dissected tissues before and after MoTissU-ExM, and highlight some of the key host and parasite structures that can be visualised following MoTissU-ExM. Finally, we provide a point-by-point protocol for how to perform MoTissU-ExM, along with details on how best to image the expanded tissues, and how to troubleshoot common issues.

cell biology↗

bioRxiv2023

Anopheles salivary apyrase regulates blood meal hemostasis and drives malaria parasite transmission

Mosquito salivary proteins play a crucial role in regulating hemostatic responses at the bite site during blood feeding. In this study, we investigate the function of Anopheles gambiae salivary apyrase (AgApyrase) in Plasmodium transmission. Our results demonstrate that salivary apyrase interacts with and activates tissue plasminogen activator, facilitating the conversion of plasminogen to plasmin, a human protein previously shown to be required for Plasmodium transmission. Microscopy imaging shows that mosquitoes ingest a substantial amount of apyrase during blood feeding which reduces coagulation in the blood meal by enhancing fibrin degradation and inhibiting platelet aggregation. Supplementation of Plasmodium infected blood with apyrase significantly enhanced Plasmodium infection in the mosquito midgut. In contrast, AgApyrase immunization inhibited Plasmodium mosquito infection and sporozoite transmission. This study highlights a pivotal role for mosquito salivary apyrase for regulation of hemostasis in the mosquito blood meal and for Plasmodium transmission to mosquitoes and to the mammal host, underscoring the potential for new strategies to prevent malaria transmission.

microbiology↗

bioRxiv2020

A high affinity human monoclonal antibody against Pfs230 binds multiple parasite stages and blocks oocyst formation in mosquitoes

Malaria elimination requires tools that interrupt parasite transmission. Here, we characterized B cell receptor responses among Malian adults vaccinated against the first domain of the cysteine-rich 230kDa gamete surface protein Pfs2301-3 to neutralize sexual stage P. falciparum parasites and halt their further spread. We generated nine Pfs230 human monoclonal antibodies (mAbs). One mAb potently blocked transmission to mosquitoes in a complement-dependent manner and reacted strongly to gamete surface while eight mAbs showed only low or no blocking activity. This study provides a rational basis to improve malaria vaccines and develop therapeutic antibodies for malaria elimination.

immunology↗