Allobaculum mucilyticum sp. nov. and Allobaculum fili sp. nov., isolated from the human intestinal tract.

As part of a culturomics study to identify bacterial species associated with inflammatory bowel disease, a large collection of bacteria was isolated from patients with ulcerative colitis. Two of these isolates were tentatively identified as members of the family Erysipelotrichaceae. Following phylogenetic analysis based on 16S rRNA gene sequence and genome sequences, both strain 128T and 539T were found to be most closely related to Allobaculum stercoricanis, with G+C contents of 48.6 and 50.5 mol%, respectively, and the genome sizes of 2 864 314 and 2 580 362 base pairs, respectively. Strains 128T and 539T were strict anaerobe rods that grew in long chains between 37 and 42 °C. Scanning electron microscopy did not reveal flagella, fimbriae or visible endospores. Biochemical analysis showed nearly identical results for both strains with enzymatic activity of C4 and C8 esterases, acid phosphatase, naphthol-AS-BI-phosphohydrolase, ß-glucuronidase, N-acetyl-ß-glucosaminidase and arginine arylamidase. In addition, both strains produced indole and reduced nitrate. Major fatty acids were identified as C18:1 ω9c (oleic acid, 64.06% in 128T and 74.35% in 539T), C18:1 ω7c/C18:1 ω9t/C18:1 ω12t/UN17.834 (16.18 % in 128T and 6.22% in 539T) and C16:0 (6.23% in 128T and 7.37% in 538T). Based on these analyses two novel species are proposed, Allobaculum mucilyticum sp. nov. with the type strain 128T (=NCTC 14626T=DSM 112815T) and Allobaculum fili sp. nov. with the type strain 539T (=NCTC 14627T=DSM 112814T).

Identification of Allobaculum mucolyticum as a novel human intestinal mucin degrader.

The human gut microbiota plays a central role in intestinal health and disease. Yet, many of its bacterial constituents are functionally still largely unexplored. A crucial prerequisite for bacterial survival and proliferation is the creation and/or exploitation of an own niche. For many bacterial species that are linked to human disease, the inner mucus layer was found to be an important niche. Allobaculum mucolyticum is a newly identified, IBD-associated species that is thought be closely associated with the host epithelium. To explore how this bacterium is able to effectively colonize this niche, we screened its genome for factors that may contribute to mucosal colonization. Up to 60 genes encoding putative Carbohydrate Active Enzymes (CAZymes) were identified in the genome of A. mucolyticum. Mass spectrometry revealed 49 CAZymes of which 26 were significantly enriched in its secretome. Functional assays demonstrated the presence of CAZyme activity in A. mucolyticum conditioned medium, degradation of human mucin O-glycans, and utilization of liberated non-terminal monosaccharides for bacterial growth. The results support a model in which sialidases and fucosidases remove terminal O-glycan sugars enabling subsequent degradation and utilization of carbohydrates for A. mucolyticum growth. A. mucolyticum CAZyme secretion may thus facilitate bacterial colonization and degradation of the mucus layer and may pose an interesting target for future therapeutic intervention.

Behçet's Disease Under Microbiotic Surveillance? A Combined Analysis of Two Cohorts of Behçet's Disease Patients.

Background: In Behçet's disease (BD), an auto-inflammatory vasculitis, an unbalanced gut microbiota can contribute to pro-inflammatory reactions. In separate studies, distinct pro- and anti-inflammatory bacteria associated with BD have been identified. Methods: To establish disease-associated determinants, we performed gut microbiome profiling in BD patients from the Netherlands (n = 19) and Italy (n = 13), matched healthy controls (HC) from the Netherlands (n = 17) and Italy (n = 15) and oral microbiome profiling in Dutch BD patients (n = 18) and HC (n = 15) by 16S rRNA gene sequencing. In addition, we used fecal IgA-SEQ analysis to identify specific IgA coated bacterial taxa in Dutch BD patients (n = 13) and HC (n = 8). Results: In BD stool samples alpha-diversity was conserved, whereas beta-diversity analysis showed no clustering based on disease, but a significant segregation by country of origin. Yet, a significant decrease of unclassified Barnesiellaceae and Lachnospira genera was associated with BD patients compared to HC. Subdivided by country, the Italian cohort displays a significant decrease of unclassified Barnesiellaceae and Lachnospira genera, in the Dutch cohort this decrease is only a trend. Increased IgA-coating of Bifidobacterium spp., Dorea spp. and Ruminococcus bromii species was found in stool from BD patients. Moreover, oral Dutch BD microbiome displayed increased abundance of Spirochaetaceae and Dethiosulfovibrionaceae families. Conclusions: BD patients show decreased fecal abundance of Barnesiellaceae and Lachnospira and increased oral abundance of Spirochaetaceae and Dethiosulfovibrionaceae. In addition, increased fecal IgA coating of Bifidobacterium, Ruminococcus bromii and Dorea may reflect retention of anti-inflammatory species and neutralization of pathosymbionts in BD, respectively. Additional studies are warranted to relate intestinal microbes with the significance of ethnicity, diet, medication and response with distinct pro- and inflammatory pathways in BD patients.

The E3 Ubiquitin Ligase TMEM129 Is a Tri-Spanning Transmembrane Protein.

Misfolded proteins from the endoplasmic reticulum (ER) are transported back into the cytosol for degradation via the ubiquitin-proteasome system. The human cytomegalovirus protein US11 hijacks this ER-associated protein degradation (ERAD) pathway to downregulate human leukocyte antigen (HLA) class I molecules in virus-infected cells, thereby evading elimination by cytotoxic T-lymphocytes. Recently, we identified the E3 ubiquitin ligase transmembrane protein 129 (TMEM129) as a key player in this process, where interference with TMEM129 activity in human cells completely abrogates US11-mediated class I degradation. Here, we set out to further characterize TMEM129. We show that TMEM129 is a non-glycosylated protein containing a non-cleaved signal anchor sequence. By glycosylation scanning mutagenesis, we show that TMEM129 is a tri-spanning ER-membrane protein that adopts an Nexo-Ccyto orientation. This insertion in the ER membrane positions the C-terminal really interesting new gene (RING) domain of TMEM129 in the cytosol, making it available to catalyze ubiquitination reactions that are required for cytosolic degradation of secretory proteins.