HLA Alleles Associated With Risk of Ankylosing Spondylitis and Rheumatoid Arthritis Influence the Gut Microbiome.

OBJECTIVE: HLA alleles affect susceptibility to more than 100 diseases, but the mechanisms that account for these genotype-disease associations are largely unknown. HLA alleles strongly influence predisposition to ankylosing spondylitis (AS) and rheumatoid arthritis (RA). Both AS and RA patients have discrete intestinal and fecal microbiome signatures. Whether these changes are the cause or consequence of the diseases themselves is unclear. To distinguish these possibilities, we examined the effect of HLA-B27 and HLA-DRB1 RA risk alleles on the composition of the intestinal microbiome in healthy individuals. METHODS: Five hundred sixty-eight stool and biopsy samples from 6 intestinal sites were collected from 107 healthy unrelated subjects, and stool samples were collected from 696 twin pairs from the TwinsUK cohort. Microbiome profiling was performed using sequencing of the 16S ribosomal RNA bacterial marker gene. All subjects were genotyped using the Illumina CoreExome SNP microarray, and HLA genotypes were imputed from these data. RESULTS: Associations were observed between the overall microbial composition and both the HLA-B27 genotype and the HLA-DRB1 RA risk allele (P = 0.0002 and P = 0.00001, respectively). These associations were replicated using the stool samples from the TwinsUK cohort (P = 0.023 and P = 0.033, respectively). CONCLUSION: This study shows that the changes in intestinal microbiome composition seen in AS and RA are at least partially due to effects of HLA-B27 and HLA-DRB1 on the gut microbiome. These findings support the hypothesis that HLA alleles operate to cause or increase the risk of these diseases through interaction with the intestinal microbiome and suggest that therapies targeting the microbiome may be effective in preventing or treating these diseases.

Correction to: Type 1 diabetes susceptibility alleles are associated with distinct alterations in the gut microbiota.

Following publication of the original article [1] it came to the attention of the Production Editor that Figs. 1 and 2 had not been replaced with the newly revised figures supplied by the authors (the originals being unusable due to poor quality image and text).

Type 1 diabetes susceptibility alleles are associated with distinct alterations in the gut microbiota.

BACKGROUND: Dysbiosis of the gut microbiota has been implicated in the pathogenesis of many autoimmune conditions including type 1 diabetes (T1D). It is unknown whether changes in the gut microbiota observed in T1D are due to environmental drivers, genetic risk factors, or both. Here, we have performed an analysis of associations between the gut microbiota and T1D genetic risk using the non-obese diabetic (NOD) mouse model of T1D and the TwinsUK cohort. RESULTS: Through the analysis of five separate colonies of T1D susceptible NOD mice, we identified similarities in NOD microbiome that were independent of animal facility. Introduction of disease protective alleles at the Idd3 and Idd5 loci (IL2, Ctla4, Slc11a1, and Acadl) resulted in significant alterations in the NOD microbiome. Disease-protected strains exhibited a restoration of immune regulatory pathways within the gut which could also be reestablished using IL-2 therapy. Increased T1D disease risk from IL-2 pathway loci in the TwinsUK cohort of human subjects resulted in some similar microbiota changes to those observed in the NOD mouse. CONCLUSIONS: These findings demonstrate for the first time that type 1 diabetes-associated genetic variants that restore immune tolerance to islet antigens also result in functional changes in the gut immune system and resultant changes in the microbiota.

Genetic association of ankylosing spondylitis with TBX21 influences T-bet and pro-inflammatory cytokine expression in humans and SKG mice as a model of spondyloarthritis.

OBJECTIVES: Ankylosing spondylitis (AS) is a highly heritable immune-mediated arthropathy. Inflammation in AS is poorly understood. TBX21 encodes T-bet, a transcription factor, lying within a locus with genome-wide significant association with AS. T-bet is implicated in innate and adaptive immunity. However, the role of T-bet in AS pathogenesis is unclear. METHODS: We assessed the importance of T-bet in disease development and progression in peripheral blood mononuclear cells from 172 AS cases and 83 healthy controls carrying either risk or protective alleles of the peak AS-associated TBX21 single nucleotide polymorphism. Kinetics and localisation of T-bet expression in the SKG mouse model of spondyloarthropathy was examined, along with the impact of Tbx21 knockout on arthritis development in SKG mice. RESULTS: Patients with AS had higher T-bet expression than healthy individuals, driven predominantly by natural killer and CD8+ T cells, with expression levels in CD8+ T cells completely distinguishing AS cases from healthy controls. T-bet expression was increased in AS cases carrying risk compared with protective alleles of rs11657479. In curdlan-treated SKG mice, T-bet expression increased early after disease initiation and persisted throughout the course of disease. There was marked reduction in gut and peripheral joint inflammation, and less IFNγ-producing and IL-17-producing CD8+ T cells, in Tbx21-/- compared with wild-type SKG mice. CONCLUSIONS: AS-associated variants in TBX21 influence T-bet expression. T-bet+ innate and adaptive immune cells have altered IL-17 and IFNγ, and early activation marker CD69 expression than T-bet cells. This indicates that T-bet is a major component of inflammatory pathways of spondyloarthropathy in humans and mice.

Functional Genomics and Its Bench-to-Bedside Translation Pertaining to the Identified Susceptibility Alleles and Loci in Ankylosing Spondylitis.

Ankylosing spondylitis (AS) is a highly heritable disease for which there is a great unmet need for improved therapies. Genetics research has identified several major pathways involved in the disease, from which treatments have either now entered clinical practice or are in development. In particular, therapies targeting the IL-23 pathway were repositioned for use in AS following the discovery of multiple genes in the pathway as determinants of AS risk. Discovery of the association of aminopeptidase genes with AS, and subsequently with psoriasis, inflammatory bowel disease and other conditions, has triggered research into therapies targeting this pathway. The AS-genetic associations point to involvement of gut mucosal immunity in driving disease, and metagenomic studies have provided strong support that AS is a disease driven by interaction between the gut microbiome and host immune system, providing a rationale for the exploration of gut-targeted therapies for the disease.

MixMC: A Multivariate Statistical Framework to Gain Insight into Microbial Communities.

Culture independent techniques, such as shotgun metagenomics and 16S rRNA amplicon sequencing have dramatically changed the way we can examine microbial communities. Recently, changes in microbial community structure and dynamics have been associated with a growing list of human diseases. The identification and comparison of bacteria driving those changes requires the development of sound statistical tools, especially if microbial biomarkers are to be used in a clinical setting. We present mixMC, a novel multivariate data analysis framework for metagenomic biomarker discovery. mixMC accounts for the compositional nature of 16S data and enables detection of subtle differences when high inter-subject variability is present due to microbial sampling performed repeatedly on the same subjects, but in multiple habitats. Through data dimension reduction the multivariate methods provide insightful graphical visualisations to characterise each type of environment in a detailed manner. We applied mixMC to 16S microbiome studies focusing on multiple body sites in healthy individuals, compared our results with existing statistical tools and illustrated added value of using multivariate methodologies to fully characterise and compare microbial communities.

The intestinal microbiome in human disease and how it relates to arthritis and spondyloarthritis.

Humans and microbes have developed a symbiotic relationship over time, and alterations in this symbiotic relationship have been linked to several immune mediated diseases such as inflammatory bowel disease, type 1 diabetes and spondyloarthropathies. Improvements in sequencing technologies, coupled with a renaissance in 16S rRNA gene based community profiling, have enabled the characterization of microbiomes throughout the body including the gut. Improved characterization and understanding of the human gut microbiome means the gut flora is progressively being explored as a target for novel therapies including probiotics and faecal microbiota transplants. These innovative therapies are increasingly used for patients with debilitating conditions where conventional treatments have failed. This review discusses the current understanding of the interplay between host genetics and the gut microbiome in the pathogenesis of spondyloarthropathies, and how this may relate to potential therapies for these conditions.

Ascites bacterial burden and immune cell profile are associated with poor clinical outcomes in the absence of overt infection.

Bacterial infections, most commonly spontaneous bacterial peritonitis in patients with ascites, occur in one third of admitted patients with cirrhosis, and account for a 4-fold increase in mortality. Bacteria are isolated from less than 40% of ascites infections by culture, necessitating empirical antibiotic treatment, but culture-independent studies suggest bacteria are commonly present, even in the absence of overt infection. Widespread detection of low levels of bacteria in ascites, in the absence of peritonitis, suggests immune impairment may contribute to higher susceptibility to infection in cirrhotic patients. However, little is known about the role of ascites leukocyte composition and function in this context. We determined ascites bacterial composition by quantitative PCR and 16S rRNA gene sequencing in 25 patients with culture-negative, non-neutrocytic ascites, and compared microbiological data with ascites and peripheral blood leukocyte composition and phenotype. Bacterial DNA was detected in ascitic fluid from 23 of 25 patients, with significant positive correlations between bacterial DNA levels and poor 6-month clinical outcomes (death, readmission). Ascites leukocyte composition was variable, but dominated by macrophages or T lymphocytes, with lower numbers of B lymphocytes and natural killer cells. Consistent with the hypothesis that impaired innate immunity contributes to susceptibility to infection, high bacterial DNA burden was associated with reduced major histocompatibility complex class II expression on ascites (but not peripheral blood) monocytes/macrophages. These data indicate an association between the presence of ascites bacterial DNA and early death and readmission in patients with decompensated cirrhosis. They further suggest that impairment of innate immunity contributes to increased bacterial translocation, risk of peritonitis, or both.

Brief Report: Intestinal Dysbiosis in Ankylosing Spondylitis.

OBJECTIVE: Ankylosing spondylitis (AS) is a common, highly heritable immune-mediated arthropathy that occurs in genetically susceptible individuals exposed to an unknown but likely ubiquitous environmental trigger. There is a close relationship between the gut and spondyloarthritis, as exemplified in patients with reactive arthritis, in whom a typically self-limiting arthropathy follows either a gastrointestinal or urogenital infection. Microbial involvement in AS has been suggested; however, no definitive link has been established. The aim of this study was to determine whether the gut in patients with AS carries a distinct microbial signature compared with that in the gut of healthy control subjects. METHODS: Microbial profiles for terminal ileum biopsy specimens obtained from patients with recent-onset tumor necrosis factor antagonist-naive AS and from healthy control subjects were generated using culture-independent 16S ribosomal RNA gene sequencing and analysis techniques. RESULTS: Our results showed that the terminal ileum microbial communities in patients with AS differ significantly (P < 0.001) from those in healthy control subjects, driven by a higher abundance of 5 families of bacteria (Lachnospiraceae [P = 0.001], Ruminococcaceae [P = 0.012], Rikenellaceae [P = 0.004], Porphyromonadaceae [P = 0.001], and Bacteroidaceae [P = 0.001]) and a decrease in the abundance of 2 families of bacteria (Veillonellaceae [P = 0.01] and Prevotellaceae [P = 0.004]). CONCLUSION: We show evidence for a discrete microbial signature in the terminal ileum of patients with AS compared with healthy control subjects. The microbial composition was demonstrated to correlate with disease status, and greater differences were observed between disease groups than within disease groups. These results are consistent with the hypothesis that genes associated with AS act, at least in part, through effects on the gut microbiome.

Does the microbiome play a causal role in spondyloarthritis?

The purpose of this study is to review the potential causal role of the microbiome in the pathogenesis of spondyloarthritis. The method used for the study is literature review. The microbiome plays a major role in educating the immune response. The microbiome is strongly implicated in inflammatory bowel disease which has clinical and genetic overlap with spondyloarthritis. The microbiome also plays a causal role in bowel and joint disease in HLA B27/human beta 2 microglobulin transgenic rats. The mechanism(s) by which HLA B27 could influence the microbiome is unknown but theories include an immune response gene selectivity, an effect on dendritic cell function, or a mucosal immunodeficiency. Bacteria are strongly implicated in the pathogenesis of spondyloarthritis. Studies to understand how HLA B27 affects bacterial ecosystems should be encouraged.

Microbes, the gut and ankylosing spondylitis.

It is increasingly clear that the interaction between host and microbiome profoundly affects health. There are 10 times more bacteria in and on our bodies than the total of our own cells, and the human intestine contains approximately 100 trillion bacteria. Interrogation of microbial communities by using classic microbiology techniques offers a very restricted view of these communities, allowing us to see only what we can grow in isolation. However, recent advances in sequencing technologies have greatly facilitated systematic and comprehensive studies of the role of the microbiome in human health and disease. Comprehensive understanding of our microbiome will enhance understanding of disease pathogenesis, which in turn may lead to rationally targeted therapy for a number of conditions, including autoimmunity.