Cindy Boer

Microbiome Composition, Joint Pain and Inflammation | 237 5.2 Table 3: Results of the association analysis of Streptococcus and knee joint effusion Taxonomy N Model 1 CoE Model 1 P-value Model 2 CoE Model 2 P-value Class Bacilli 314 9.4 x10 -03 3.4 x10 -02 2.7 x10 -03 3.5 x10 -01 Order Lactobacillales 314 9.8 x10 -03 2.7 x10 -02 2.7 x10 -03 3.6 x10 -01 Family Streptococcaceae 310 9.6 x10 -03 1.7 x10 -02 3.0 x10- 03 2.6 x10 -01 Genus Streptococcus 308 1.0 x10 -02 1.3 x10 -02 3.3 x10- 03 2.1 x10 -01 Knee joint inflammation was measured as severity of effusion as measured on knee MRI. Knee MRI’s were only available for an all-female obese subgroup of the Rotterdam Study Microbiome dataset (n=373). First model assessed the association of Knee effusion with the microbiome, adjusted for age, sex, DNA isolation batch and TimeInMail (technical covariates). Second model was WOMAC-pain score adjusted for age, sex, technical covariates and, effusion severity. P-values were determined by MaAsLin analysis. N=number of individuals in cohort where microbial abundancy is not zero for that taxonomy. CoE coefficient Independent replication of Streptococcus association We sought replication for all four associations with WOMAC pain, i.e., class, order, family and, the bacterial genus of Streptococcus, in an independent Dutch cohort, Life- Lines-DEEP (LLD) ( Supplementary Table 7 ). LLD has a lower sample size (n=867), younger age (mean age=45.6) and fewer individuals with OA-related knee pain (WOM- AC pain>0, n=197), however, the average kneeWOMAC painwas very similar compared with the Rotterdam Study cohort (RS=0.9 and LLD=0.9, Table 1 and Supplementary Table 7 ). Despite lower power in the replication cohort we observed a significant as- sociation (p-value<0.05) between knee WOMAC-pain scores and all four taxonomies to the genus of Streptococcus (coefficient replication =3.3×10 −03 , p-value replication =3.7×10 −02 , MaAsLin, Table 2 ). Also, in the meta-analysis of RS and LLD, greater Streptococcus spp. abundance was significantly associated with higher knee WOMAC pain (p-value- meta =1.3x10 −08 , MaAsLin, n=2256, Table 2 ). After adjusting for BMI, we found signifi- cant replication on class, order and the family level of Streptococcacea (coefficient rep- lication =2.7×10 −03 , p-value replication =3.5×10 −02 , MaAsLin, Supplementary Table 8 ). In the replication study, additional adjustment for BMI slightly attenuated the association, a 9.1% decrease in coefficient, for the genus of Streptococcus ( Supplementary Table 8 ). This was in concordance with the 14.5% decrease in coefficient for streptococcus, seen in the discovery cohort after BMI adjustment ( Supplementary Table 2 ). In the BMI adjusted meta-analysis for RS and LLD, the association between knee WOMAC pain and Streptococcus spp. was highly significant (p-value meta =1.1×10 −06 , METAL, n=2,256, Sup- plementary Table 8 ).