SEMI-AUTOMATED DETERMINATION OF TOXOPLASMA GONDII LOCALIZATION IN THE CENTRAL NERVOUS SYSTEM

Abstract

I have studied the interactions between cells of the brain and the neurotropic parasite Toxoplasma gondii. Using an in vivo system that allows us to permanently mark and identify brain cells injected with Toxoplasma protein, we have identified that Toxoplasma-injected neurons (TINs) are heterogeneously distributed throughout the brain. Unfortunately, standard methods to quantify and map heterogeneous cell populations onto a reference brain atlas are time consuming and prone to user bias. We developed a novel MATLAB-based semi-automated quantification and mapping program to allow the rapid and consistent mapping of heterogeneously distributed cells on to the Allen Institute Brain Atlas. The procedure takes advantage of simple immunohistochemistry labeling techniques, use of a standard microscope with a motorized stage, and low cost computing that can be readily obtained at most research institutes. The system uses a two-threshold background subtraction to identify and quantify cells of interest, and automated transformation to align experimental images onto a brain atlas. We demonstrate that we reliably quantify and neuroanatomically localize TINs with low intra- or inter-observer variability. Additionally, we show that specific regions of the mouse brain are enriched with TINs after T. gondii infection. The discovery of TIN enrichment in specific brain regions was facilitated by our quantification method described here, which allowed reliable visualization, quantification, and mapping of TINs populations in immunolabeled sections across whole mouse brains.