iiinova — where your digital self keeps evolving

bishara marzook @ghostpathogen.bsky.social · 27/04/2026

Hi all, I'd like to share some news - I will soon be moving to Lisbon, Portugal to start my very own lab @gimminstitute.bsky.social! The Epithelial Biology of Apicomplexan Infections lab (Epi-Api lab for short) will focus on the special relationship between #epithelial cells and #Cryptosporidium.

Picture of Bishara standing by a coastline at daytime, with a blue-green sea and palm trees in the background. She is holding a pastel de nata (Portuguese custard tart) and smiling.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

We tested lapaquistat on a mouse model of Cryptosporidium infection and discovered that it did a pretty good job at restricting overall parasite burdens, and reducing numbers of visible parasites in the ileum. 16/

Graphs showing parasite burdens in mice treated with two doses of lapaquistat or vehicle only controls, showing lower overall burdens in drug-treated mice.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

We made transgenic parasites to test how important those remaining glutathione-recycling genes were. We can induce loss of glutathione reductase by adding rapamycin to induce DNA excision. Parasites with induced loss of this gene did not survive in mice, showing glutathione recycling is essential.

Graph showing burdens of transgenic parasites in mice given either DMSO or Rap - short for rapamycin. Parasites in mice treated with Rapamycin did not establish a productive infection and fell below the detection threshold.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

Glutathione is one of life's ways of dealing with oxidative damage, it's a crucial metabolite across all kingdoms of life. But of course, Crypto being crypto, has lost the genes to make its own glutathione, unlike other apicomplexan parasites and most eukaryotes. But it can still use and recycle it.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

We found that accumulated squalene can act as an anti-oxidant in epithelial cells. Our KO cell line that accumulates squalene had lower levels of reactive oxygen species (ROS) compared to parental controls. Plus, adding a common anti-oxidant like N-Ac boosts parasites to similar levels.

Violin plot comparing mean fluorescent intensities of CellROX in wild-type cells compared to SQLE knockout cells, showing lower levels in SQLE knockouts.

Immunofluorescence images showing number of parasites, and female parasites in particular under different drug treatments. Lapaquistat inhibits squalene synthesis and reduces visible parasites, while NB-598 enhances squalene accumulation which enhances parasite growth, similar to the antioxidant N-acetylcysteine, or N-Ac.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

We reproduced this phenotype using chemical inhibitors of either gene (growth is either floored or enhanced) or by creating a KO cell line that accumulates squalene in host cells. This was especially true for development of parasite sexual stages which seem to teeter on the metabolite squalene. 9/

Violin plot showing the percentage of female parasites, using female-specific antibodies against proteins COWP1 or DMC1, is inhibited by Lapaquistat, called Lap in the graph, or enhanced by NB-598.

Violin plot showing enhanced growth of parasites in SQLE-KO cells that accumulate squalene.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

Amazingly, knocking out two consecutive genes in the human cholesterol synthesis pathway either inhibited or promoted parasite growth! This seemed to hinge on an accumulation of the intermediary metabolite squalene that is sandwiched in between these two genes. 8/

A host epithelial cell houses the intracellular parasite Cryptosporidium. The host cholesterol biosynthesis pathway, going from acetyl-coA to cholesterol is drawn within the host cell. The intermediary metabolite Squalene is highlighted in the middle of the pathway, in between to genes FDFT1 and SQLE. Inhibiting FDFT1 inhibits parasite growth, while inhibiting SQLE enhances parasite growth.

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bishara marzook @ghostpathogen.bsky.social · 06/12/2024

Have you ever wondered how knocking out every single gene in the human genome, one by one, might affect your intracellular pathogen of interest? We did, & boy was it a wild ride! Happy to share our preprint discovering the essential host genome for Cryptosporidium - www.biorxiv.org/content/10.1...

Schematic showing the experimental outline for a full genome arrayed CRISPR screen. 384-well plates containing cells with an arrayed crRNA library are infected with Cryptosporidium parasites, which undergo asexual rounds of replication intracellularly, followed by sexual development to male and female forms. Plates are then fixed and immunolabeled to detect all parasites, female parasites, host actin recruited to parasite vacuoles, and host nuclei. Three z-stacks were imaged by high content microscopy to detect apically located parasites as well as host cell actin and nuclei.

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bishara marzook @ghostpathogen.bsky.social · 04/12/2023

Indole-3-carbinol (I3C) is a phytochemical enriched in Brassica veggies. It activates AHR. Supplementing mouse diets with I3C augments their IEL population. So we fed mice I3C and challenged them with Cryptosporidium. They were protected! I went straight home and ate some kale

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bishara marzook @ghostpathogen.bsky.social · 04/12/2023

Amazingly, transferring IELs to immunodeficient mice was enough to rescue them from infection-related morbidity and they had much lower parasite burdens. Go go barrier cytotoxic activity! But what about the indoles? Here's where we got really excited -

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bishara marzook @ghostpathogen.bsky.social · 04/12/2023

We then narrowed this down to CD8+ intraepithelial lymphocytes (IELs). These cells highly express AHR, depend on the presence of AHR ligands, and proliferate in response to Crypto infections.

A bar graph showing that all subsets of CD8 positive cells proliferate - using Ki67 as a marker - in response to a Cryptosporidium infection.

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bishara marzook @ghostpathogen.bsky.social · 04/12/2023

We decided to dig deeper. If AHR (& its activating ligands) can protect against Crypto, how? Using a series of transgenic mouse models, we found that AHR expression - particularly in immune cells - was important to curb Crypto infections.

Graph comparing Cryptosporidium parasite burdens in mice - wildtype mice versus those with AHR expression lost in immune cells. Graph shows that the Cryptosporidium burden is higher in mice without immune cell-AHR expression.

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