Mitchell, Gabriel, Torres, Laura, Fishbaugher, Matthew, Lam, Melanie, Chuenchob, Eve, Zalpuri, Reena, Ramasubban, Shreya, Baxter, Caitlin, Flannery, Erika, Harupa-Chung, Anke, Mikolajczak, Sebastian and Jorgens, Danielle (2024) Correlative light-electron microscopy methods to characterize the ultrastructural features of the replicative and dormant liver stages of malaria. Malaria journal, 23. p. 53. ISSN 1475-2875

Abstract

Background:
The infection of the liver is an obligatory step leading to malaria disease. Following hepatocyte invasion, malaria parasites either differentiate into liver stage schizonts or hypnozoites, which can lie dormant for extended periods of time before reactivating and causing relapses. The liver stages of malaria remain elusive because of technical challenges hindering access to well-established technologies, including electron microscopy. A deeper understanding of hypnozoite biology could prove essential in the development of radical cure therapeutics against malaria.

Results:
The liver stages of the non-relapsing rodent parasite Plasmodium berghei and the relapsing simian parasite Plasmodium cynomolgi were characterized in human Huh7 cells or primary monkey hepatocytes using Correlative Light-Electron Microscopy (CLEM). Specifically, CLEM approaches that rely on GFP-expressing P. berghei (GFP-CLEM) or on immunofluorescence assays (IFA-CLEM) for both P. berghei and P. cynomolgi were used for imaging of liver stages. The results from P. berghei showed that host and parasite organelles can be identified and imaged at high resolution using both CLEM approaches. However, while IFA-CLEM was associated with more pronounced extraction of cellular content, samples’ features were generally well preserved. Using IFA-CLEM, a collection of micrographs was acquired for P. cynomolgi liver stage schizonts and hypnozoites, demonstrating the potential of this approach as a useful tool for characterizing the liver stages of relapsing malaria species.

Conclusions:
We developed a CLEM protocol for imaging the liver stages of malaria species that are difficult to study due to a lack of established genetic systems. This study also provides a dataset that characterizes the ultrastructural features of liver stage schizonts and hypnozoites from the relapsing simian malaria species P. cynomolgi.

Item Type:	Article
Date Deposited:	09 Mar 2024 00:46
Last Modified:	09 Mar 2024 00:46
URI:	https://oak.novartis.com/id/eprint/52093