Assessing whether rectal swabs reflect appendiceal microbiota profiles in acute appendicitis: a 16S rRNA-based...

Acute appendicitis is associated with characteristic changes in the intestinal microbiota, but direct sampling of appendiceal contents is invasive and cannot be performed in healthy controls. We therefore evaluated whether rectal swabs could partially capture appendiceal microbiome signatures in adults with acute appendicitis.

In a prospective cross-sectional study, we enrolled adults with acute appendicitis and healthy volunteers between October 2023 and December 2024. Four types of samples were collected: feces from healthy controls (HC), appendiceal luminal contents from patients with acute appendicitis (AC), intraoperative rectal swabs from patients with acute appendicitis (RS), and initial postoperative feces from patients with acute appendicitis (IF; first stool within 24 h after surgery). 16 S rRNA gene (V3-V4) sequencing was performed, and reads were processed with QIIME2. Alpha and beta diversity, differential taxonomic composition, and PICRUSt2-based functional predictions were compared across matrices. Genus-level and functional concordance between paired AC-RS samples was assessed.

After quality control, 64 AC, 34 RS, 24 IF, and 29 HC samples were included. Phylogenetic diversity (PD whole-tree) was higher in AC and RS than HC, with AC also higher than RS; IF showed lower PD than AC. Bray-Curtis principal coordinate analysis showed AC forming a distinct cluster separated from HC and RS along PC1, whereas IF overlapped with HC and RS. AC, RS, and IF were enriched for Escherichia/Shigella and Fusobacterium and depleted in butyrate-producing genera such as Faecalibacterium compared with HC. In the 21 paired AC-RS cases, genus-level relative abundances and several predicted functional pathways showed concordance, indicating that RS captured many but not all appendiceal dysbiosis features.

Our findings suggest that intraoperative rectal swabs may partially reflect appendiceal microbiome alterations at the genus and pathway levels and may serve as a minimally invasive adjunct for microbiome profiling in acute appendicitis. However, these associations are inferred from 16 S amplicon data in a modestly sized, antibiotic-exposed cohort and should be validated using shotgun metagenomics in larger, clinically stratified populations.