Epigenetic Lymphocyte Subset Associations Across Allergic Phenotypes in a Predominantly Pediatric Cohort

Background/Objectives: DNA methylation-based lymphocyte enumeration may provide an alternative to conventional immunophenotyping, but its associations with allergic phenotypes require careful interpretation. We examined these associations in a predominantly pediatric cohort. Methods: This retrospective study included 206 participants aged 1-57 years (189 aged <18 years; 57.8% male; median age 5 years, IQR 3-8), evaluated between 24 November 2025 and 28 February 2026. Absolute subset counts were derived from the epigenetic i.Mune TBNK assay and CBC lymphocyte counts. Group comparisons used Mann-Whitney U tests with Benjamini-Hochberg correction across 56 comparisons. Firth models adjusted for three subset counts, CD4/CD8 ratio, log(IgE+1), age, and sex; percentage-based, comorbidity-adjusted, and age-restricted sensitivity analyses were added. IgE-containing models included 201 complete cases. Results: Allergic rhinitis was associated with lower CD4+ counts in the primary model (OR per 100 cells/µL 0.923; p = 0.0009), with the association retained after comorbidity adjustment and age restriction but not in the percentage model. Atopic dermatitis was associated with log(IgE+1) (OR 1.475; p = 0.0009). Food allergy was associated with higher CD19+ counts in the primary model (OR per 100 cells/µL 1.060; p = 0.0341), but this association was not retained in sensitivity analyses. No absolute NK-count difference was found for food allergy. Single-marker discrimination was modest (CD4+ AUC for rhinitis 0.711). Conclusions: These exploratory associations are sensitive to the measurement scale and model specification. They do not establish phenotype-specific epigenetic mechanisms, standalone diagnostic utility, or functional immune monitoring.