Intracellular Parasites Toxoplasma gondii and Besnoitia besnoiti, Unveiled in Single Host Cells Using AP-SMALDI MS Imaging.

Abstract

The obligate intracellular apicomplexan parasites Toxoplasma gondii and Besnoitia besnoiti are important causes of disease in both humans and cattle. To date, effective specific treatments are lacking for both infections. To counteract severe symptoms leading to, e.g., disabilities and even abortion in the case of human toxoplasmosis and bovine besnoitiosis, novel targets are required for development of drugs and vaccines. A promising emerging technique for molecular characterization of organisms is high-resolution atmospheric-pressure scanning microprobe matrix-assisted laser desorption/ionization (AP-SMALDI) mass spectrometry imaging (MSI) which enables semiquantitative visualization of metabolite distributions. MSI was here used to trace and characterize lipid metabolites in primary bovine umbilical vein endothelial cells (BUVECs) upon infection with tachyzoites, an early and pathogenic fast-replicating life stage of T. gondii and B. besnoiti. A cell bulk, derived from noninfected controls and parasite-infected cell pellets, was analyzed by AP-SMALDI MSI in technical and biological triplicates. Multivariate statistical analysis including hierarchical clustering and principle component analysis revealed infection-specific metabolites in both positive- and negative-ion mode, identified by combining database search and LC-MS2 experiments. MSI analyses of host cell monolayers were conducted at 5 μm lateral resolution, allowing single apicomplexan-infected cells to be allocated. This is the first mass spectrometry imaging study on intracellular T. gondii and B. besnoiti infections and the first detailed metabolomic characterization of B. besnoiti tachyzoites. MSI was used here as an efficient tool to discriminate infected from noninfected cells at the single-cell level in vitro.

• Authors: Patrik Kadesch, Tobias Hollubarsch, Stefanie Gerbig, Lars Schneider, Liliana M R Silva, Carlos Hermosilla, Anja Taubert, Bernhard Spengler
• Journal: Journal of the American Society for Mass Spectrometry (J Am Soc Mass Spectrom) Vol. 31 Issue 9 Pg. 1815-1824 (Sep 02 2020) ISSN: 1879-1123 [Electronic] United States
• PMID: 32830963 (Publication Type: Journal Article)

Topics

• Animals
• Cattle
• Cells, Cultured
• Coccidiosis (diagnostic imaging, parasitology)
• Endothelial Cells (cytology, parasitology)
• Intracellular Space (parasitology)
• Molecular Imaging
• Sarcocystidae (pathogenicity)
• Single-Cell Analysis
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization (methods)
• Toxoplasma (pathogenicity)
• Toxoplasmosis, Animal (diagnostic imaging, parasitology)