Some are more equal than others : the role of ‘keystone’ species in the degradation of recalcitrant substrates

Peer reviewedPublisher PD

The presence of the putative Gardnerella vaginalis sialidase A gene in vaginal specimens is associated with bacterial vaginosis biofilm

Bacterial vaginosis (BV) is a difficult-to-treat recurrent condition in which health-associated lactobacilli are outnumbered by other anaerobic bacteria, such as Gardnerella vaginalis. Certain genotypes of G. vaginalis can produce sialidase, while others cannot. Sialidase is known to facilitate the destruction of the protective mucus layer on the vaginal epithelium by hydrolysis of sialic acid on the glycans of mucous membranes. This process possibly facilitates adhesion of bacterial cells on the epithelium since it has been linked with the development of biofilm in other pathogenic conditions. Although it has not been demonstrated yet, it is probable that G. vaginalis benefits from this mechanism by attaching to the vaginal epithelium to initiate biofilm development. In this study, using vaginal specimens of 120 women enrolled in the Ring Plus study, we assessed the association between the putative G. vaginalis sialidase A gene by quantitative polymerase chain reaction (qPCR), the diagnosis of BV according to Nugent score, and the occurrence of a BV-associated biofilm dominated by G. vaginalis by fluorescence in situ hybridisation (FISH). We detected the putative sialidase A gene in 75% of the G. vaginalis-positive vaginal specimens and found a strong association (p<0.001) between the presence of a G. vaginalis biofilm, the diagnosis of BV according to Nugent and the detection of high loads of the G. vaginalis sialidase A gene in the vaginal specimens. These results could redefine diagnosis of BV, and in addition might guide research for new treatment

Development of Functional Microfold (M) Cells from Intestinal Stem Cells in Primary Human Enteroids.

Background &amp; aimsIntestinal microfold (M) cells are specialized epithelial cells that act as gatekeepers of luminal antigens in the intestinal tract. They play a critical role in the intestinal mucosal immune response through transport of viruses, bacteria and other particles and antigens across the epithelium to immune cells within Peyer's patch regions and other mucosal sites. Recent studies in mice have demonstrated that M cells are generated from Lgr5+ intestinal stem cells (ISCs), and that infection with Salmonella enterica serovar Typhimurium increases M cell formation. However, it is not known whether and how these findings apply to primary human small intestinal epithelium propagated in an in vitro setting.MethodsHuman intestinal crypts were grown as monolayers with growth factors and treated with recombinant RANKL, and assessed for mRNA transcripts, immunofluorescence and uptake of microparticles and S. Typhimurium.ResultsFunctional M cells were generated by short-term culture of freshly isolated human intestinal crypts in a dose- and time-dependent fashion. RANKL stimulation of the monolayer cultures caused dramatic induction of the M cell-specific markers, SPIB, and Glycoprotein-2 (GP2) in a process primed by canonical WNT signaling. Confocal microscopy demonstrated a pseudopod phenotype of GP2-positive M cells that preferentially take up microparticles. Furthermore, infection of the M cell-enriched cultures with the M cell-tropic enteric pathogen, S. Typhimurium, led to preferential association of the bacteria with M cells, particularly at lower inoculum sizes. Larger inocula caused rapid induction of M cells.ConclusionsHuman intestinal crypts containing ISCs can be cultured and differentiate into an epithelial layer with functional M cells with characteristic morphological and functional properties. This study is the first to demonstrate that M cells can be induced to form from primary human intestinal epithelium, and that S. Typhimurium preferentially infect these cells in an in vitro setting. We anticipate that this model can be used to generate large numbers of M cells for further functional studies of these key cells of intestinal immune induction and their impact on controlling enteric pathogens and the intestinal microbiome

Sustainable Power Production in a Membrane-less and Mediator-less Wastewater Microbial Fuel Cell

Microbial fuel cells (MFCs) fed with wastewater are currently considered a feasible strategy for production of renewable electricity at low cost. A membrane-less MFC with biological cathode was built from a compact wastewater treatment reactor. When operated with an external resistance of 250 Ohm, the MFC produced a long-term power of approximately 70 mW/m2 for ten months. Denaturing Gradient Gel Electrophoresis (DGGE) analysis of the cathode biomass when the MFC was closed on a 2100 Ohm external resistance showed that the sequenced bands were affiliated with Firmicutes, -Proteobacteria, -Proteobacteria, -Proteobacteria, and Bacteroidetes groups. When the external resistance was varied between 250 and 2100 Ohm, sustainable resistance decreased from 900 to 750 Ohm, while sustainable power output decreased from 32 to 28 mW/m2. It is likely that these effects were caused by changes in the microbial ecology of anodic and cathodic biomass attached to the electrodes. Results suggest that cathodic biomass enrichment in “electroactive” bacteria may improve MFCs power output in a similar fashion to what has been already observed for anodic biomass

The OX-44 molecule couples to signaling pathways and is associated with CD2 on rat T lymphocytes and a natural killer cell line.

The MRC OX-44 molecule, which is expressed on all peripheral leukocytes, identifies the subset of thymocytes capable of proliferating in response to alloantigens and lectins (Paterson, D.J., J.R. Green, W.A. Jefferies, M. Puklavec, and A.F. Williams. 1987. J. Exp. Med. 165:1). When we isolated monoclonal antibodies (mAbs) on the basis of their ability to activate the phosphatidylinositol signaling pathway in RNK-16 cells (a rat leukemia line with natural killer activity), three of the resulting mAbs recognized the OX-44 molecule. Addition of these mAbs to RNK-16 elicits protein tyrosine phosphorylation, generates inositol phosphates, and increases the concentration of cytoplasmic free calcium. These responses require the addition of intact mAb and are not observed with F(ab')2 fragments. One of these mAbs (7D2) is mitogenic for freshly isolated rat splenic T cells and synergizes with a mAb to the T cell antigen receptor in this activation. A 50-60-kD glycoprotein coprecipitates with the OX-44 molecule from RNK-16 cells and rat splenic T cells. Peptide mapping and reprecipitation studies indicate that the coprecipitating molecule is CD2. Thus, the OX-44 molecule can couple to multiple signaling pathways and associates with CD2 on both RNK-16 and rat T cells

Predominant variable region gene usage by gamma/delta T cell receptor-bearing cells in the adult thymus.

Previous studies have indicated that the diversity of gamma genes expressed by gamma/delta-bearing murine T cells is limited, but comparable information concerning the expressed diversity of delta genes is lacking. In this study, we have investigated the rearrangement and expression of delta and gamma genes in T cell hybridomas that express gamma/delta T cell receptors. Three productive delta chain cDNA clones were isolated (delta 7.3, delta 7.1, and delta 2.3) that encode new variable region sequences. Two of the delta cDNAs differ significantly from those observed in the V alpha repertoire. In addition, one cDNA expressed a new J delta region (J delta 2), which was localized between J delta 1 and C delta genes. Using these and other delta gene probes and gamma gene probes, we found that five independent hybridomas expressed four different V delta s and three different V gamma s. However, analysis of an enriched population of gamma/delta-expressing cells from the adult thymus suggests that only a few V delta genes and one V gamma gene are used by the majority of the cells. These results suggest that important components of receptor chain that contribute to specificity (i.e., the germline V gene sequences) are relatively nondiverse in the thymic gamma/delta population

Glycomimicry: Display of the GM3 sugar epitope on Escherichia coli and Salmonella enterica sv Typhimurium

Oligosaccharides present on the surface of pathogenic bacteria play an important role in their interaction with their host. Bacteria with altered cell surface structures can be used to study these interactions, and glycoengineering represents a tool to display a glycoepitope on a different bacterium. Here, we present non-pathogenic Escherichia coli and Salmonella enterica serovar Typhimurium expressing the sialyllactose oligosaccharide epitope of the ganglioside GM3. By expression of the galactosyltransferase LgtE and the sialic acid transferase Lst as well as the CMP-sialic acid synthetase SiaB from Neisseria gonorrhoeae and Neisseria meningitidis in engineered strains devoid of the sialic acid catabolism, the GM3 sugar epitope was displayed on these bacteria as demonstrated by live cell immunostaining and a detailed analysis of their lipooligosaccharides. These strains offer the possibility to investigate the role of sialic acid in the recognition of bacteria by the immune system in a non-pathogenic backgroun