ProbeST: a custom probe design pipeline for dual host-pathogen Spatial Transcriptomics

Area of Science:

• Genomics
• Bioinformatics
• Molecular Biology

Background:

• Probe-based Spatial Transcriptomics (ST) offers spatially-resolved transcriptome profiling.
• Current commercial probe sets limit ST applications primarily to human and mouse studies.
• A need exists for adaptable methods to apply ST to diverse organisms.

Purpose of the Study:

• To develop an open-source computational pipeline, ProbeST, for designing custom gene-specific probe sets.
• To enable the application of probe-based ST to non-model organisms.
• To facilitate studies of host-pathogen interactions using ST.

Main Methods:

• ProbeST pipeline development for custom probe set design.
• Validation using Formalin-fixed paraffin-embedded (FFPE) mouse enteroid models infected with Salmonella Typhimurium.
• Simultaneous detection of host and pathogen transcripts using custom and existing probe panels.

Main Results:

• ProbeST successfully designed custom pathogen probes for Salmonella Typhimurium.
• Simultaneous detection of host (Mefv, Tnf, Anxa1) and pathogen transcripts was achieved.
• High probe specificity and low sensitivity were observed, with identified host genes colocalizing with pathogen genes.

Conclusions:

• The ProbeST workflow is reproducible and expands probe-based ST capabilities.
• This computational tool democratizes ST for non-model organisms.
• ProbeST facilitates detailed spatial analysis of host-pathogen interactions.