Molecular Characterization of Circulating Microbiome Signatures in Rheumatoid Arthritis

Abstract

Rheumatoid Arthritis (RA) has been increasingly associated with perturbations to the microbial communities that reside in and on the body (the microbiome), in both human and animal studies. To date, such studies have mainly focused on the microbial communities that inhabit the gut and oral cavity. Mounting evidence suggests that microbial DNA can be detected in the blood circulation using a range of molecular methods. This DNA may represent an untapped pool of biomarkers that have the potential to report on changes to the microbiome of distant sites (e.g., example, the gut and oral cavity). To this end, through amplification and sequencing of the bacterial 16S rRNA variable region four, we evaluated the presence and identity of microbial DNA in blood samples obtained from RA patients (both prior to and 3 months following the instigation of treatment) in comparison to a small number of healthy control subjects and samples obtained from patients with ankylosing spondylitis (AS) and psoriatic arthritis (PA). Bacterial-derived DNA was identified in the majority of our patient samples. Taxonomic classification revealed that the microbiome community in RA was distinct from AS, PA, and the healthy state. Through analysis of paired patient samples obtained prior to and 3 months following treatment (V0 vs. V3), we found the microbiome to be modulated by treatment, and in many cases, this shift reduced the distance between these samples and the healthy control samples, suggesting a partial normalization following treatment in some patients. This effect was especially evident in seronegative arthritis patients. Herein, we provide further evidence for the existence of a blood microbiome in health and identify specific taxa modulated in disease and following treatment. These blood-derived signatures may have significant utility as disease biomarkers and suggest this area warrants further investigation.

Comparison of Blood Bacterial Communities in Periodontal Health and Periodontal Disease

“…Most bacteraemia and blood metagenomic studies employ methods such as PCI-extraction of blood fractions or systems such as the Phusion Blood Direct kit (Thermo Fisher Scientific), both of which generate total DNA (Amar et al, 2013;Rajendhran et al, 2013; Dinakaran et al, 2014;Lelouvier et al, 2016;Paisse et al, 2016;Gosiewski et al, 2017;Li et al, 2018;Castillo et al, 2019;Hammad et al, 2019). These studies tend to show Proteobacteria as the dominant phylum found in blood, but with high variability between studies at lower taxonomic levels, even taking into account diseaseassociated dysbiosis.…”

“…Streptococcal species of the viridans group, containing the Streptococcus mitis (S. mitis) group are the taxa most commonly associated with community-acquired native valve infective endocarditis. Periodontitis, oral pathogens and the dysbiotic oral microbiome have also been linked to the aetiology of other inflammatory diseases such as rheumatoid arthritis (Loyola-Rodriguez et al, 2010;Zhang et al, 2015;Konig et al, 2016;Beyer et al, 2018;Correa et al, 2019;Gomez-Banuelos et al, 2019;Hammad et al, 2019), late onset Alzheimer's disease (Singhrao and Olsen, 2019) and type II diabetes, which has been linked to distinct microbiomes in the gut (Wellen and Hotamisligil, 2005;Larsen et al, 2010), blood (Qiu et al, 2019), and oral cavity (Long et al, 2017).…”

“…One study showed Actinobacteria as dominant in health, but, by contrast Proteobacteria was dominant in disease (Dinakaran et al, 2014). A circulating microbiome study of rheumatoid arthritis (RA) subjects (Hammad et al, 2019) showed that all samples (healthy controls and RA) contained 45.8% Proteobacteria and 31% Firmicutes. Paisse et al (2016) suggested that high levels of bacterial-derived DNA, measured at 4 X 10 7 16S rRNA gene sequences (corresponding to 10 6 -10 7 bacterial genomes) per ml, are present in blood as cell-free or immune processed fragments.…”

Circulating 16S RNA in Biofluids: Extracellular Vesicles as Mirrors of Human Microbiome?

“…More recently, a manuscript from Hammad and co-authors described the profiles of circulating microbial DNA in patients with rheumatoid arthritis (RA) in comparison with patients affected by ankylosing spondylitis (AS) or psoriatic arthritis (PA) and healthy control subjects [ 46 ]. Bacterial community members were identified by sequencing of the 16S rRNA variable region 4 in all samples.…”

Table_1.xlsx

“…See Supplementary Figure 3. Dinakaran et al, 2014;Lelouvier et al, 2016;Paisse et al, 2016;Gosiewski et al, 2017;Li et al, 2018;Castillo et al, 2019;Hammad et al, 2019). These studies tend to show Proteobacteria as the dominant phylum found in blood, but with high variability between studies at lower taxonomic levels, even taking into account diseaseassociated dysbiosis.…”