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The colonisation of specific body sites in contact with the external environment by microorganisms is both well-described and universally accepted, whereas, the existence of microbial evidence in other “classically sterile” locations including the blood, synovial space, and lungs, is a relatively new concept. Increasingly, a role for the microbiome in disease is being considered, and it is therefore necessary to increase our understanding of these. To date, little data support the existence of a “synovial fluid microbiome”.   Methods  The presence and identity of bacterial and fungal DNA in the synovial fluid of rheumatoid arthritis (RA) patients and healthy control subjects was investigated through amplification and sequencing of the bacterial 16S rRNA gene and fungal internal transcribed spacer region 2 respectively. Synovial fluid concentrations of the cytokines IL-6, IL-17A, IL22 and IL-23 were determined by ELISA.    Results  Bacterial 16S rRNA genes were detected in 87.5% RA patients, and all healthy control subjects. At the phylum level, the microbiome was predominated by  Proteobacteria  (Control = 83.5%, RA = 79.3%) and  Firmicutes  (Control = 16.1%, RA = 20.3%), and to a much lesser extent,  Actinobacteria  (Control = 0.2%, RA = 0.3%) and  Bacteroidetes  (Control = 0.1%, RA = 0.1%). Fungal DNA was identified in 75% RA samples, and 88.8% healthy controls. At the phylum level, synovial fluid was predominated by members of the Basidiomycota (Control = 53.9%, RA = 46.9%) and Ascomycota (Control = 35.1%, RA = 50.8%) phyla. Statistical analysis revealed key taxa that were differentially present or abundant dependent on disease status.    Conclusions  This study reports the presence of a synovial fluid microbiome, and determines that this is modulated by disease status (RA) as are other classical microbiome niches.

Molecular characterisation of the synovial fluid microbiome in rheumatoid arthritis patients and healthy control subjects