Fareha Umam

Background HLA-B27 is a susceptibility factor for enthesitis-related arthritis. HLA-B27 transgenic rats do not develop disease in a germ-free state, suggesting involvement of the gut in the pathogenesis of the disease. There is limited data on the effect of HLA-B27 carriage and gut flora in JIA-ERA. Methods Stool samples from 10 HLA-B27-positive and 5 HLA-B27-negative JIA-ERA patients, and 10 HLA-B27-negative healthy controls were collected after consent. Fecal calprotectin and serum LPS levels were measured using ELISA. The fecal flora was assessed using 16S rRNA based sequencing. Results HLA-B27-positive patients had higher fecal calprotectin levels (p < 0.001) compared to HLA-B27-negative patients and HC. However, serum LPS levels were not different. Alpha and beta diversity of fecal flora were similar in HLA-B27-positive and negative patients, but different compared to HC. HLA-B27-positive patients had higher abundance of family Veillonellaceae, Prevotellaceae and genera Moryella, Intestinimonas, Fournierella, Leyella, Massiliprevotella, Segatella, and Bilophila and lower abundance of genera Bifidobacterium compared to HC. At the species level, the abundance of Coprococcus eutactus and Prevotella copri was higher while that of Dorea longicatena was lower in HLA-B27-positive patients compared to HC. In addition, there was lower abundance of the genera Fusicatenibacter and Lachnospiraceae NC2004 group, species belonging to genera Agathobacter, Fusicatenibacter, Butyricicoccus, and Acutalibacter and higher abundance of genera Leyella in HLA-B27-positive patients compared to HLA-B27-negative patients. Conclusion Lower abundance of anti-inflammatory and higher abundance of pro-inflammatory bacteria in HLA-B27-positive JIA-ERA patients by inducing gut inflammation may play a role in disease pathogenesis. Key Points • The presence of HLA-B27 in JIA-ERA patients is associated with a decline in the population of beneficial bacteria with anti-inflammatory properties and an increase in gut inflammation. • Exploration of supplementation with butyrate or SCFA to improve gut health may be beneficial in patients with JIA-ERA.

For an effective control of tuberculosis (TB), there is a persistent need for biomarkers that can report true estimates of TB infection (TBI) and predict its progression towards active TB disease. We investigated whether the cell‐mediated immune responses to Mycobacterium tuberculosis (Mtb) antigens could provide such biomarkers. The study subjects (n = 174) comprised a cohort of smear‐positive, drug‐sensitive, HIV‐negative pulmonary TB patients (n = 54) and their household contacts (HC, n = 120). Whole blood cultures, in the presence or absence of Mtb antigens‐ membrane (MtM), purified protein derivative (PPD) and alpha‐crystallin (Acr), or the mitogen PHA were subjected to determinations, by flow cytometry, for T cell proliferative and, by ELISA, for IFN‐γ, TNF‐α, and IL‐6 cytokine responses. Additionally, serum levels of the three cytokines were also estimated. The strongest cell‐proliferative and cytokine responses were induced by MtM and IL‐6 was the most abundantly produced cytokine. While none of the responses induced by Mtb antigens or the serum cytokines levels could discriminate between TB and HC, the ex vivo cytokine responses induced by PHA or ‘spontaneously’ could apparently do so. The concentrations of IFN‐γ induced by PHA in TB blood cultures were significantly lower than in HC cultures (AUC = 0.72). Conversely, the spontaneous IFN‐γ or TNF‐α secretions in TB cultures were significantly higher than in HC cultures (AUC = 0.66). Our results suggest that IL‐6 responses to MtM could be a sensitive indicator of TBI, and low levels of PHA‐induced or high levels of spontaneous IFN‐γ secretions in HC blood cultures may indicate a progressive infection.

This study was aimed at identifying the B cell responses which could distinguish between ‘latent tuberculosis infection (LTBI)’ and active TB disease. Study subjects were smear-positive TB patients (n = 54) and their disease-free household contacts (HHCs, n = 120). The sera were used for determination of antibody levels (ΔOD values) against Mycobacterium tuberculosis membrane (MtM) antigens by ELISA and for visualisation of seroreactive MtM antigens by immunoblotting. B cell subsets in whole blood samples were determined by flow cytometry. In TB sera, levels of IgG antibodies were significantly higher than IgM and IgA whereas IgM and IgA antibody levels were comparable. Conversely, HHC sera had significantly higher IgM antibody levels than IgG and IgA. The ratio of IgM to IgG antibodies in HHCs were also significantly higher than in patients. Immunoblotting revealed that some of the MtM antigens (<10, ~12 and ~25 kDa) reacted with TB as well as HHC sera whereas some other antigens (~16, ~36, ~45 and ~60 kDa) reacted with most of TB and a subset of HHC sera. Frequencies of classical memory B cells (cMBCs, CD19+CD27+) were significantly higher, and of IgG+ cMBCs were significantly lower in HHCs than in patients. Frequencies of IgA+ cMBCs in HHCs and patients were comparable but both were significantly higher than the corresponding frequencies of IgG+ cMBCs. Frequencies of IgA+ atypical MBCs (aMBCs, CD19+CD27-) in HHCs and patients were also comparable and significantly higher than the IgG+ aMBCs. The plasmablast (CD19+CD27++CD38++) frequencies in HHCs and patients were comparable. These results suggest that the IgM/IgG antibody ratio, antibody binding to selected MtM antigens and relative frequencies of MBC subsets could indicate protective or pathogenic immune responses following the primary infection with Mtb. Responses that orchestrate protection leading to a ‘quiescent’ LTBI may provide clues to an effective vaccination strategy against TB.