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Abstract While triple-negative breast cancer (TNBC) is known to be heterogeneous at the genomic and transcriptomic levels, spatial information on tumor organization and cell composition is still lacking. Here, we investigate TNBC tumor architecture including its microenvironment using spatial transcriptomics on a series of 92 patients. We perform an in-depth characterization of tumor and stroma organization and composition using an integrative approach combining histomorphological and spatial transcriptomics. Furthermore, a detailed molecular characterization of tertiary lymphoid structures leads to identify a gene signature strongly associated to disease outcome and response to immunotherapy in several tumor types beyond TNBC. A stepwise clustering analysis identifies nine TNBC spatial archetypes, further validated in external datasets. Several spatial archetypes are associated with disease outcome and characterized by potentially actionable features. In this work, we provide a comprehensive insight into the complexity of TNBC ecosystem with potential clinical relevance, opening avenues for treatment tailoring including immunotherapy.

Abstract Upon infecting its vertebrate host, the malaria parasite initially invades the liver where it undergoes massive replication, whilst remaining clinically silent. The coordination of host responses across the complex liver tissue during malaria infection remains unexplored. Here, we perform spatial transcriptomics in combination with single-nuclei RNA sequencing over multiple time points to delineate host-pathogen interactions across Plasmodium berghei-infected liver tissues. Our data reveals significant changes in spatial gene expression in the malaria-infected tissues. These include changes related to lipid metabolism in the proximity to sites of Plasmodium infection, distinct inflammation programs between lobular zones, and regions with enrichment of different inflammatory cells, which we term ‘inflammatory hotspots’. We also observe significant upregulation of genes involved in inflammation in the control liver tissues of mice injected with mosquito salivary gland components. However, this response is considerably delayed compared to that observed in P. berghei-infected mice. Our study establishes a benchmark for investigating transcriptome changes during host-parasite interactions in tissues, it provides informative insights regarding in vivo study design linked to infection and offers a useful tool for the discovery and validation of de novo intervention strategies aimed at malaria liver stage infection.

Abstract Several important human infectious diseases are caused by microscale-sized parasitic nematodes like filarial worms. Filarial worms have their own spatial tissue organization; to uncover this tissue structure, we need methods that can spatially resolve these miniature specimens. Most filarial worms evolved a mutualistic association with endosymbiotic bacteria Wolbachia. However, the mechanisms underlying the dependency of filarial worms on the fitness of these bacteria remain unknown. As Wolbachia is essential for the development, reproduction, and survival of filarial worms, we spatially explored how Wolbachia interacts with the worm’s reproductive system by performing a spatial characterization using Spatial Transcriptomics (ST) across a posterior region containing reproductive tissue and developing embryos of adult female Brugia malayi worms. We provide a proof-of-concept for miniature-ST to explore spatial gene expression patterns in small sample types, demonstrating the method’s ability to uncover nuanced tissue region expression patterns, observe the spatial localization of key B. malayi - Wolbachia pathway genes, and co-localize the B. malayi spatial transcriptome in Wolbachia tissue regions, also under antibiotic treatment. We envision our approach will open up new avenues for the study of infectious diseases caused by micro-scale parasitic worms.

Global transcriptional differences across lobular units in the liver remain unknown. Here the authors perform spatial transcriptomics of liver tissue to delineate transcriptional differences in physical space, confirm lobular zonation along transcriptional gradients and suggest the presence of previously uncharacterized structures within liver tissue.