Summary: This paper presents MBRarefy, an R package that provides a reproducible workflow for alpha diversity analysis under confounding from heterogeneous library sizes. Building on the multi-bin rarefying approach in Li et al (2024), MBRarefy supports alpha diversity association analysis with repeated rarefying, bin-wise testing, and cross-bin meta-analysis. A key new feature is automated, data-adaptive selection of library size bin thresholds via a genetic algorithm (GA), which replaces ad hoc cutpoints with an objective optimization procedure based on the rarefying-derived profiles. The package also supports routine data-management tasks, including file-based sample-wise processing and standardized output generation, enabling users to execute the full analysis pipeline from raw count files to combined inferential results. Availability and implementation: The R package MBRarefy is freely available on GitHub at https://github.com/mli171/MBRarefy.

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