Maximum depth sequencing reveals an ON/OFF replication slippage switch and apparent in vivo selection for bifidobacterial pilus expression

The human gut microbiome, of which the genus Bifidobacterium is a prevalent and abundant member, is thought to sustain and enhance human health. Several surface-exposed structures, including so-called sortase-dependent pili, represent important bifidobacterial gut colonization factors. Here we show that expression of two sortase-dependent pilus clusters of the prototype Bifidobacterium breve UCC2003 depends on replication slippage at an intragenic G-tract, equivalents of which are present in various members of the Bifidobacterium genus. The nature and extent of this slippage is modulated by the host environment. Involvement of such sortase-dependent pilus clusters in microbe-host interactions, including bacterial attachment to the gut epithelial cells, has been shown previously and is corroborated here for one case. Using a Maximum Depth Sequencing strategy aimed at excluding PCR and sequencing errors introduced by DNA polymerase reagents, specific G-tract sequences in B. breve UCC2003 reveal a range of G-tract lengths whose plasticity within the population is functionally utilized. Interestingly, replication slippage is shown to be modulated under in vivo conditions in a murine model. This in vivo modulation causes an enrichment of a G-tract length which appears to allow biosynthesis of these sortase-dependent pili. This work provides the first example of productive replication slippage influenced by in vivo conditions. It highlights the potential for microdiversity generation in “beneficial” gut commensals

Ursodeoxycholic acid and lithocholic acid exert anti-inflammatory actions in the colon

Inflammatory bowel diseases (IBD) comprise a group of common and debilitating chronic intestinal disorders for which currently available therapies are often unsatisfactory. The naturally occurring secondary bile acid, ursodeoxycholic acid (UDCA), has well-established anti-inflammatory and cytoprotective actions and may therefore be effective in treating IBD. We aimed to investigate regulation of colonic inflammatory responses by UDCA and to determine the potential impact of bacterial metabolism on its therapeutic actions. The anti-inflammatory efficacy of UDCA, a nonmetabolizable analog, 6 alpha-methyl-UDCA (6-MUDCA), and its primary colonic metabolite lithocholic acid (LCA) was assessed in the murine dextran sodium sulfate (DSS) model of mucosal injury. The effects of bile acids on cytokine (TNF-alpha, IL-6, Il-1 beta, and IFN-alpha) release from cultured colonic epithelial cells and mouse colonic tissue in vivo were investigated. Luminal bile acids were measured by gas chromatography-mass spectrometry. UDCA attenuated release of proinflammatory cytokines from colonic epithelial cells in vitro and was protective against the development of colonic inflammation in vivo. In contrast, although 6-MUDCA mimicked the effects of UDCA on epithelial cytokine release in vitro, it was ineffective in preventing inflammation in the DSS model. In UDCA-treated mice, LCA became the most common colonic bile acid. Finally, LCA treatment more potently inhibited epithelial cytokine release and protected against DSS-induced mucosal inflammation than did UDCA. These studies identify a new role for the primary metabolite of UDCA, LCA, in preventing colonic inflammation and suggest that microbial metabolism of UDCA is necessary for the full expression of its protective actions.NEW & NOTEWORTHY On the basis of its cytoprotective and anti-inflammatory actions, the secondary bile acid ursodeoxycholic acid (UDCA) has well-established uses in both traditional and Western medicine. We identify a new role for the primary metabolite of UDCA, lithocholic acid, as a potent inhibitor of intestinal inflammatory responses, and we present data to suggest that microbial metab-olism of UDCA is necessary for the full expression of its protective effects against colonic inflammation

The influence of soft contact lens wear and two weeks cessation of lens wear on corneal curvature.

INTRODUCTION: Accurate corneal measurements are crucial in corneal refractive surgery (CRS) to ensure successful outcomes. Soft contact lens (SCL) wear may result in changes to corneal curvature and structure. United States Food and Drug Administration (FDA) pre-operative guidelines recommend that prior to CRS, SCL wearers cease SCL wear for "at least two weeks before examination and treatment" [1]. Corneal curvature changes induced by SCL wear may take longer than two weeks to resolve. PURPOSE: To examine the effect of SCL wear on corneal curvature before and following two weeks SCL wear cessation. To explore the possible impact of different SCL materials and years of SCL wear. METHODS: Retrospective data analysis, between a group of SCL wearers (SCL: n=45); and a non-contact lens control group (NCL: n=45). Corneal curvature parameters were measured using the Pentacam (Oculus, Germany), before and following two weeks cessation of SCL wear. RESULTS: No significant differences in keratometry or Sagittal radius of curvature between SCL and NCL groups prior to or following SCL cessation. Tangential radius of curvature showed significant inferior steepening for the SCL group prior to SCL cessation (SCL vs. NCL; 7.77±0.30mm vs. 7.90±0.30mm; p=0.04). Following two weeks cessation of SCL wear this appeared to have resolved. CONCLUSIONS: Two weeks cessation of SCL wear appears sufficient for resolution of corneal curvature changes with modern SCL materials and years of SCL wear. However, further studies with longer lens deprivation periods are required to ensure stability for all SCL wearing patients

Characterization of AQPs in mouse, rat and human colon and their selective regulation by bile acids

In normal individuals, the epithelium of the colon absorbs 1.5-2 L of water a day to generate dehydrated feces. However, in the condition of bile acid malabsorption (BAM), an excess of bile acids in the colon results in diarrhea. Several studies have attempted to address the mechanisms contributing to BAM induced by various bile acids. However, none have addressed a potential dysregulation of aquaporin water channels, which are responsible for the majority of transcellular water transport in epithelial cells, as a contributing factor to the onset of diarrhea and the pathogenesis of BAM. In this study we aimed to systematically analyze the expression of AQPs in colonic epithelia from rat, mouse and human and determine whether their expression is altered in a rat model of BAM. Mass spectrometry-based proteomics, RT-PCR, and western blotting identified various AQPs in isolated colonic epithelial cells from rats (AQP1, 3, 4, 7, 8) and mice (AQP1, 4, 8). Several AQPs were also detected in human colon (AQP1, 3, 4, 7-9). Immunohistochemistry localized AQP1 to the apical plasma membrane of epithelial cells in the bottom of the crypts, whereas AQP3 (rat, human) and AQP4 (mice, human) were localized predominantly in the basolateral plasma membrane. AQP8 was localized intracellularly and at the apical plasma membrane of epithelial cells. Rats fed sodium cholate for 72 h had significantly increased fecal water content, suggesting development of BAM associated diarrhea. Colonic epithelial cells isolated from this model had significantly altered levels of AQP3, 7, and 8, suggesting that these AQPs may be involved in the pathogenesis of bile acid induced diarrhea

UnThree – Dimensionally ordered macroporous amorphous C/TiO2 composite electrodes for lithium-ion batteries

A facile method utilizing colloidal templating and sucrose as a carbon precursor is used to synthesize highly ordered, porous inverse opal structures as C/TiO2 nanocomposites. Material characterization shows amorphous TiO2 and a large pore size of ∼400 nm allowing for enhanced electrolyte penetration. C/TiO2 inverse opals materials as electrodes in Li-ion battery half cells demonstrate discharge and charge capacities of ∼870 mAh g−1 and 470 mAh g−1 , respectively, at a current density of 150 mA g−1 The enhanced capacities, which surpass theoretical limits for TiO2 and carbon based on intercalation reactions, are analyzed under voltammetric conditions to assess relative contributions to capacity from diffusion-limited intercalation and capacitive charge compensation reactions. The porous structure contributes to excellent capacity retention, rate performance and improved Coulombic efficiency (99.6% after 250 cycles), compared to individual carbon and TiO2 inverse opals. </p

Comparing cycling and rate response of SnO₂ macroporous anodes in lithium-ion and sodium-ion batteries

Tin oxide (SnO2) is a useful anode material due to its high capacity (1493 mAh g−1 and 1378 mAh g−1 vs Li/Li+ and vs Na/Na+, respectively) and natural abundance (tin is one of the thirty most abundant elements on Earth). Unfortunately, only moderate electrical conductivity and significant volume expansion of up to 300% for Li-ion, and as much as 520% for Na-ion can occur. Here, we use an ordered macroporous interconnected inverse opal (IO) architectures to enhance rate capability, structural integrity, and gravimetric capacity, without conductive additives and binders. Excellent capacity retention is shown during cycling vs Na/Na+ relative to Li/Li+. Cyclic voltammetry (CV) analysis, galvanostatic cycling, and differential capacity analysis extracted from rate performance testing evidence the irreversibility of the oxidation of metallic Sn to SnO2 during charge. This behavior allows for a very stable electrode during cycling at various rates. A stable voltage profile and rate performance is demonstrated for both systems. In a Na-ion half cell, the SnO2 retained >76% capacity after 100 cycles, and a similar retention after rate testing.</p

Maximum depth sequencing reveals an ON/OFF replication slippage switch and apparent in vivo selection for bifdobacterial pilus expression

The human gut microbiome, of which the genus Bifidobacterium is a prevalent and abundant member, is thought to sustain and enhance human health. Several surface-exposed structures, including so-called sortase-dependent pili, represent important bifidobacterial gut colonization factors. Here we show that expression of two sortase-dependent pilus clusters of the prototype Bifidobacterium breve UCC2003 depends on replication slippage at an intragenic G-tract, equivalents of which are present in various members of the Bifidobacterium genus. The nature and extent of this slippage is modulated by the host environment. Involvement of such sortase-dependent pilus clusters in microbe-host interactions, including bacterial attachment to the gut epithelial cells, has been shown previously and is corroborated here for one case. Using a Maximum Depth Sequencing strategy aimed at excluding PCR and sequencing errors introduced by DNA polymerase reagents, specific G-tract sequences in B. breve UCC2003 reveal a range of G-tract lengths whose plasticity within the population is functionally utilized. Interestingly, replication slippage is shown to be modulated under in vivo conditions in a murine model. This in vivo modulation causes an enrichment of a G-tract length which appears to allow biosynthesis of these sortase-dependent pili. This work provides the first example of productive replication slippage influenced by in vivo conditions. It highlights the potential for microdiversity generation in “beneficial” gut commensals.</p