Malaria is a mosquito-borne disease that infects millions of people every year and kills hundreds of thousands of those infected. The Plasmodium parasites are the cause of malaria and are transmitted through the bite of an infected Anopheles mosquito.
A malaria research study from Pascini et. al at the National Institute of Allergy and Infectious Diseases (NIAID) highlights the possibility of expressing other human proteins from the fibrinolytic system to prevent the transmission of malaria from other parasite species. Due to the reduced chance of selective pressure against the modifications, this study also provides a novel engineered mosquito line that could be used to aid in the mitigation of malaria in high risk areas.
As part of the validation of the novel mosquito models, Western Blot analysis to detect expression of the transgenic human PAI-1 protein was used. The authors used the Azure Imager c600 to acquire these data. In addition to chemiluminescence, the Azure c600 also detects near infrared, white light and more.
Pascini et al. engineered a transgenic Anopheles mosquito that constitutively expresses human PAI-1 in the midgut and/or salivary glands to target plasminogen activation on the surface of the Plasmodium parasite. While the parasite can still bind plasminogen, the ability of the parasite to activate plasminogen in either area is blocked due to the expression of PAI-1. Since the parasite makes use of plasminogen to invade and progress through its life cycle, the authors accurately predicted that presence of PAI-1 would severely impair malaria transmission.
The authors investigated how the expression of human PAI-1 would affect the mosquitoes and found architectural changes to the salivary glands that ultimately resulted in fewer Plasmodium sporozoites in the region. Even in the midgut, they saw reduced Plasmodium infection. Reduced numbers still left the possibility for malaria transmission, as only a few sporozoites are required for infection.
Expression of human PAI-1 strongly impaired transmission of malaria between hosts. This marks the first study to report a transgenic mosquito that strongly impairs Plasmodium survival through the expression of a human protein.
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Research surrounding malaria transmission
While there have been many efforts to prevent malaria transmission, research is challenging due to the complex life cycle of the parasite. However, various efforts have been made to hinder the ability of mosquitoes to carry the parasite through genetic modifications. These have shown great promise, but have not provided an ultimate solution. Usually, this genetic modification approach has focused on two main areas: suppressing or eradicating the mosquito population or modifying the mosquito population’s ability to successfully carry or transmit the parasite itself.
While initially effective, selective pressure eventually creates a work around. To this end, Pascini et. al (2022) reported recently that they created a transgenic mosquito that did not directly target the parasite or the mosquito, but could still impair malaria transmission. This was accomplished through utilizing the mammalian fibrinolytic system, which is important for the parasitic life cycle. Because this approach does not alter the parasite directly, the risk for selective pressure is greatly reduced.
How mosquitos contract Plasmodium
To select a target that would affect the Plasmodium but not require changes to the parasite itself, the authors looked at outside factors involved in the life cycle of Plasmodium.
Mosquitos contract Plasmodium gametocytes through feeding on infected organisms, including humans. Upon ingestion by the mosquito, these gametocytes travel into the midgut of the mosquito where they are fertilized and form an oocyst . The oocyst then releases thousands of sporozoites which invade the mosquito’s salivary glands. The infected mosquito releases these sporozoites into the bloodstreams of any humans from which it takes future blood meals. The sporozoites eventually form gametocytes. When another mosquito takes a blood meal from a newly infected human, the cycle continues.
In both the mosquito and the human, the parasite must be able to penetrate physical barriers such as the fibrin network, extracellular matrices, and cellular barriers. Previously, Pascini and colleagues showed that some of these physical barriers were overcome by way of the mammalian fibrinolytic system.
Fibrinolysis occurs when plasminogen is cleaved and becomes the protease plasmin, which in turn degrades fibrin, a protein found in the blood that is involved in blood clotting. This activation of plasminogen into plasmin occurs through proteolytic cleavage by tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA). This process is regulated by the protease inhibitor plasminogen activator inhibitor 1 (PAI-1), which inhibits tPA and uPA.
Plasmodium makes use of this fibrinolytic system to infiltrate its host. Both tPA and uPA are used to activate plasminogen, while plasmin helps break down the extracellular matrix, improving the parasite’s motility within the host.
- Pascini, T.V., Jeong, Y.J., Huang, W. et al. Transgenic Anopheles mosquitoes expressing human PAI-1 impair malaria transmission. Nat Commun 13, 2949 (2022). https://doi.org/10.1038/s41467-022-30606-y