Describing the Urinary Microbiome in Pre-operative Urine Specimens of Lichen Sclerosus Induced and Non-Lichen Sclerosus Induced Urethral Stricture Disease
Amanda Sherman, MD1, Travis Sullivan, MS1, Harjivan Kohli, MD1, Eric Burks, MD2, Kimberly Rieger-Christ, PhD1, Alex Vanni, MD1.
1Lahey Hospital and Medical Center, Burlington, MA, USA, 2Boston Medical Center, Boston, MA, USA.
Background: Lichen sclerosus (LS) is a chronic inflammatory condition that can affect both the penile skin and urethral epithelium in men. Over time, severe impairment of sexual and urinary function can result, from preputial adhesions and phimosis, skin tearing with erections, acquired buried penis, and urethral stricture disease (USD). The pathophysiology of LS USD is poorly understood, with a heterogeneous presentation and disease severity. This study seeks to describe differences in the urinary microbiome of patients with pathologically confirmed LS USD vs non-LS USD.
Methods: An IRB-approved protocol of men with USD was performed. Pre-operative clean-catch voided urine was collected in 34 men with pathologically confirmed LS USD and non-LS USD. Bacterial genomic DNA was extracted using the PowerMag Soil DNA Isolation Kit. 16S rRNA gene sequencing was performed using the MiSeq platform for paired-end sequencing. Amplicon sequence variants (ASVs) were generated via dada2 v1.16.0. ASV taxonomy was assigned up to genus level using the SILVA v.138 database. The pathologic evaluation of strictures was based on 5 typical histologic features of LS (Prabhu et al 2013). For the purpose of this study, two groups were formed based on the following LS score: non-LS strictures (LS score 0-1) and LS strictures (LS score 3-5).
Results: Sufficient bacterial DNA for analysis was obtained from 19 patients (10 non-LS USD and 9 LS USD). Significant differences in alpha diversity (within sample variance) and beta diversity (between sample variance) were observed (p<0.05) between the two cohorts. ɑ diversity, as described by observed ASV, showed median ASVs for NLS samples were 25, whereas LS were 57 (p=0.016). ẞ diversity is significantly different between scores 0 and 5 (p<0.05) by PERMANOVA analysis including Bray-Curtis, and weighted and unweighted UniFrac scores. 2 phyla were noted to be significantly different between status groups, bacteroidota more common in LS (p=0.012) and firmicutes more present in NLS (p=0.037). Overall, alpha and beta diversity of samples is statistically significant between disease status groups, with a trend towards increasing diversity with higher pathologic scores.
Conclusions: LS USD exhibits significant alterations in diversity and differential abundance of urine microbiota compared to non-LS USD controls, with a trend of diversity increasing along with pathologic LS score. While further validation in a larger sample is necessary, this finding could be used to guide further investigation into the role of bacteria and the urinary microbiome in LS USD pathogenesis, the severity of presentation, and stricture recurrence. 
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