Cindy Boer

238 | Chapter 5.2 Streptococcus association with knee joint inflammation If Streptococcus spp. abundance is causally related to higher knee WOMAC pain, a pos- sible mechanism might be through local priming of macrophages in the synovial lining resulting in inflammation[22]. For a random subset of the Rotterdam Study Microbiome cohort, knee magnetic resonance images (MRI) were available (n=373, all females)[32] at the time of microbiome measurements. Knee joint inflammation was assessed by scoring the amount of effusion in the tibiofemoral joint and in the patellofemoral joint for both knees. We found that higher WOMAC-pain scores were significantly correlated with more knee effusion (Spearman correlation r =0.14, p-value=9.0×10 −03 ). In addi- tion, we observed that greater Streptococcus spp. abundance was significantly associ- ated with more effusion in the knee joints (coefficient =1.0×10 −02 , p-value=1.3×10 −02 , MaAsLin, Table 3 ). When WOMAC pain was added to the model, the association of knee effusion with Streptococcus spp. disappears (coefficient =3.3×10 −03 , p-value=0.21, MaAsLin), indicating that the association of Streptococcus spp. with knee pain severity is driven by knee inflammation severity. Discussion Using a large, deeply phenotyped population-based cohort, we identified a significant association between greater relative and absolute Streptococcus spp. abundance and higher OA-related knee pain. These results were validated by replication in an indepen- dent cohort and by meta-analysis. Finally, we presented evidence that this association was driven by local inflammation in the joint. We observed that the intestinal microbiome β-diversity was significantly asso- ciated with knee WOMAC scores. After testing 256 taxonomies individually, we found a microbiome-wide association with knee WOMAC pain and Streptococcus spp ., where greater Streptococcus spp. relative abundance is associated with higher knee WOMAC pain. We found this association to be robust, not be caused by outlier observations, or due to the confounding effects of smoking, alcohol intake, oral medication usage or BMI[28,29]. Neither was the association an artefact of the microbiome profiles as relative fractions of the 16S rRNA sequencing, or due to possible co-linearity in the data[30,33]. Although, the results of the sensitivity analyses were in line with a true associ- ation between Streptococcus spp. and knee WOMAC pain in our cohort, validation in an independent cohort is essential. Replication of microbiome abundance, however, is difficult, because data might not be similar between studies if sample preparation and