ASPIRE: the Amplicon Sequencing Profiler for Investigating Respiratory Ecosystems

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Abstract Microbial communities inhabiting the respiratory tract contribute to health status through interactions with host physiology, immune function, and local environmental conditions. Advances in small subunit ribosomal RNA (SSU or 16S rRNA) gene amplicon sequencing enable culture-independent profiling of microbial communities as amplicon sequence variants (ASVs), revealing links between microbial dysbiosis and respiratory diseases, and the use of mass spectrometry to measure volatile organic compounds (VOCs) in exhaled breath shows emerging promise for biomarker discovery. Here we present ASPIRE, the Amplicon Sequencing Profiler for Investigating Respiratory Ecosystems, an accessible Nextflow workflow for processing, analyzing, and interpreting linked ASV-VOC data from respiratory microbiome studies. ASPIRE is designed to support scalable comparative analysis across respiratory sample types while preserving intermediate file outputs for inspection and reuse within a standardized file structure. Availability and implementation The ASPIRE workflow, quick start guide, and usage are freely available on the GitHub platform https://github.com/hallamlab/ASPIRE . ASPIRE is implemented in Nextflow with a modular design to allow for reproducibility, extensibility and scalability.

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