16S rRNA sequencing for profiling microbial communities in idiopathic granulomatous mastitis

ABSTRACT Background Idiopathic granulomatous mastitis (IGM) is a rare, chronic, and benign inflammatory breast disease with ambiguous aetiology. Contrastingly, lactational mastitis (LM) is commonly diagnosed in breastfeeding women. To investigate IGM aetiology, we profiled the microbial flora of pus and skin in IGM and LM patients. Methods IGM patients were recruited from 2018 to 2020; LM patients were recruited as controls. 16S rRNA sequencing libraries were constructed from 16S rRNA gene amplified from total DNA extracted from pus and skin swabs. Metagenomic analysis was conducted using modified microbiome abundance analysis-suite customised R-resource for paired pus and skin samples. Microbiome multivariable association analyses were performed using linear models. Results 21 IGM and 3 LM paired pus and skin samples underwent metagenomic analysis. Bray-Curtis ecological dissimilarity distance showed dissimilarity across four sample types (IGM-pus, IGM-skin, LM-pus, LM-skin; PERMANOVA, p<0.001). No characteristic dominant genus was observed across IGM samples. IGM pus samples were more diverse than corresponding IGM skin samples (Shannon and Simpson index; Wilcoxon paired signed-rank tests, p=0.022 and p=0.07). Corynebacterium kroppenstedtii , reportedly associated with IGM in literature, was higher in IGM pus samples than paired skin sample (Wilcoxon, p=0.022). Three other species and nineteen genera were statistically significant in paired IGM pus-skin comparison after antibiotic treatment adjustment, and multiple comparisons correction. Conclusions Microbial profiles are unique between IGM and LM patients. Inter-patient variability and polymicrobial IGM pus samples cannot implicate specific genus or species as an infectious cause for IGM.