Lactate cross-feeding between Bifidobacterium species and Megasphaera indica contributes to butyrate formation in the human colonic environment.

Butyrate, a physiologically active molecule, can be synthesized through metabolic interactions among colonic microorganisms. Previously, in a fermenting trial of human fecal microbiota, we observed that the butyrogenic effect positively correlated with the increasing Bifidobacterium population and an unidentified Megasphaera species. Therefore, we hypothesized that a cross-feeding phenomenon exists between Bifidobacterium and Megasphaera, where Megasphaera is the butyrate producer, and its growth relies on the metabolites generated by Bifidobacterium. To validate this hypothesis, three bacterial species (B. longum, B. pseudocatenulatum, and M. indica) were isolated from fecal cultures fermenting hydrolyzed xylan; pairwise cocultures were conducted between the Bifidobacterium and M. indica isolates; the microbial interactions were determined based on bacterial genome information, cell growth, substrate consumption, metabolite quantification, and metatranscriptomics. The results indicated that two Bifidobacterium isolates contained distinct gene clusters for xylan utilization and expressed varying substrate preferences. In contrast, M. indica alone scarcely grew on the xylose-based substrates. The growth of M. indica was significantly elevated by coculturing it with bifidobacteria, while the two Bifidobacterium species responded differently in the kinetics of cell growth and substrate consumption. Coculturing led to the depletion of lactate and increased the formation of butyrate. An RNA-seq analysis further revealed the upregulation of M. indica genes involved in the lactate utilization and butyrate formation pathways. We concluded that lactate generated by Bifidobacterium through catabolizing xylose fueled the growth of M. indica and triggered the synthesis of butyrate. Our findings demonstrated a novel cross-feeding mechanism to generate butyrate in the human colon.IMPORTANCEButyrate is an important short-chain fatty acid that is produced in the human colon through microbial fermentation. Although many butyrate-producing bacteria exhibit a limited capacity to degrade nondigestible food materials, butyrate can be formed through cross-feeding microbial metabolites, such as acetate or lactate. Previously, the literature has explicated the butyrate-forming links between Bifidobacterium and Faecalibacterium prausnitzii and between Bifidobacterium and Eubacterium rectale. In this study, we provided an alternative butyrate synthetic pathway through the interaction between Bifidobacterium and Megasphaera indica. M. indica is a species named in 2014 and is indigenous to the human intestinal tract. Scientific studies explaining the function of M. indica in the human colon are still limited. Our results show that M. indica proliferated based on the lactate generated by bifidobacteria and produced butyrate as its end metabolic product. The pathways identified here may contribute to understanding butyrate formation in the gut microbiota.

Production of a high molecular weight levan by Bacillus paralicheniformis, an industrially and agriculturally important isolate from the buffalo grass rhizosphere.

A new exopolysaccharide (EPS) producing Gram-positive bacterium was isolated from the rhizosphere of Bouteloua dactyloides (buffalo grass) and its EPS product was structurally characterized. The isolate, designated as LB1-1A, was identified as Bacillus paralicheniformis based on 16S rRNA gene sequence and phylogenetic tree analysis. The EPS produced by LB1-1A was identified as a levan, having ß(2 → 6) linked backbone with ß(2 → 1) linkages at the branch points (4.66%). The isolate LB1-1A yielded large amount (~ 42 g/l) of levan having high weight average molecular weight (Mw) of 5.517 × 107 Da. The relatively low degree of branching and high molecular weight of this levan makes B. paralicheniformis LB1-1A a promising candidate for industrial applications.

The effectiveness of mouthwash against SARS-CoV-2 infection: A review of scientific and clinical evidence.

The COVID-19 pandemic, caused by the spread of SARS-CoV-2 infection that is mainly through the airborne transmission, is a worldwide health concern. This review seeks to assess the potential effectiveness of mouthwash in reducing the oropharyngeal load of SARS-CoV-2 based on the available evidence. Articles related to mouthwash and COVID-19 in PubMed were electronically searched in July, 2021. After manually excluding articles lacking sufficient scientific evidence or validation processes, those with inaccessible online full text, those that did not test the effectiveness of mouthwash against SARS-CoV-2, and those not written in English, 17 original and 13 review articles were chosen for this review. The eligible articles revealed that the main virucidal mechanism of mouthwash was via interactions with the viral envelope. Povidone-iodine (PVP-I), cetylpyridinium chloride (CPC), and essential oils with ethanol showed virucidal effects on SARS-CoV-2 in vitro, potentially by interfering with the viral envelope. A few clinical studies demonstrated that PVP-I, CPC, hydrogen peroxide, and chlorhexidine reduced the oropharyngeal load of SARS-CoV-2. Although the available evidence is limited, mouthwash containing PVP-I or CPC shows potential for reducing the oropharyngeal load of SARS-CoV-2 and thus may present a risk-mitigation strategy for COVID-19 patients.

Characterization of the stemness and osteogenic potential of oral and sinus mucosal cells.

BACKGROUND/PURPOSE: Covering the wounds from guided bone regeneration and sinus floor elevation with oral and sinus mucosa is a fundamental criterion for success. This study aimed to verify the regeneration capability of the mucosal connective tissue stromal cells by characterizing their stemness and osteogenic potentials. METHODS: Bone marrow stromal cells (BMSCs), alveolar mucosa cells (AMCs), keratinized gingival cells (KGCs), and sinus mucosal cells (SMCs), were isolated from four Sprague-Dawley rats. The morphology and viability of the cells were investigated under a confocal microscope and by Alamar Blue. Stem cell surface markers were evaluated by flow cytometry. Expressions of pluripotent factors after initial seeding and an early osteogenic gene following 24 h of osteoinduction were evaluated by realtime PCR. Trilineage differentiation capability in long-term inductive cell culture was assessed by Alizarin Red, Alcian Blue, and Oil Red O staining. RESULTS: BMSCs and AMCs were larger cells with smaller aspect ratios relative to KGCs and SMCs, and BMSCs revealed the greatest initial viability but the slowest proliferation. More than 94% of BMSCs, AMCs, and KGCs were double-positive for CD73 and CD90. Compared with BMSCs, AMCs expressed significantly higher Oct4 but reduced Cbfa1 after initial seeding, and AMCs and SMCs expressed significantly higher Cbfa1 following 24 h of osteoinduction. In long-term inductive cell culture, osteogenesis was observed in BMSCs, AMCs, and SMCs, chondrogenesis was observed in BMSCs, AMCs, and KGCs, and adipogenesis was evident in only BMSCs. CONCLUSION: AMCs contain a high percentage of stem/progenitor cells and show differentiation capability toward osteogenic lineage.

Influence of chain length on the colonic fermentation of xylooligosaccharides.

Xylooligosaccharides (XOS) have been employed as prebiotics containing oligomers of varying sizes or molecular ratios. XOS with a low degree of polymerization (DP) has been demonstrated to have high prebiotic potential. However, there is limited information regarding the specific chain length of XOS required to elicit distinct responses in the gut microbiota. In this study, we aimed to explore whether variations in XOS DP could alter the fate of colonic fermentation. Five XOS fractions (BWXFs) with DP ranges of >40, 20-40, 10-20, 5-10, and 2-4 were prepared by beechwood xylan autohydrolysis and tested on human gut microbiota. Extracellular XOS degradation was observed for molecules with a DP exceeding 5. BWXF treatments altered the microbial community structures, and substrate size-dependent effects on the microbial composition and metabolic outputs were observed. Bacteroidaceae were specifically enriched by xylan. Lachnospiraceae were particularly stimulated by XOS with a DP of 20-40 and 2-4. Bifidobacteriaceae were notably enriched by XOS with a DP of 5-20. High butyrate yields were obtained from cultures containing long-chain BWXFs. Microbiota responses differed with XOS DP composition changes, and microbial competition with XOS with a DP of 2-4 requires further exploration.

Algorithm-Driven Tele-otoscope for Remote Care for Patients With Otitis Media.

OBJECTIVE: The COVID-19 pandemic has spurred a growing demand for telemedicine. Artificial intelligence and image processing systems with wireless transmission functionalities can facilitate remote care for otitis media (OM). Accordingly, this study developed and validated an algorithm-driven tele-otoscope system equipped with Wi-Fi transmission and a cloud-based automatic OM diagnostic algorithm. STUDY DESIGN: Prospective, cross-sectional, diagnostic study. SETTING: Tertiary Academic Medical Center. METHODS: We designed a tele-otoscope (Otiscan, SyncVision Technology Corp) equipped with digital imaging and processing modules, Wi-Fi transmission capabilities, and an automatic OM diagnostic algorithm. A total of 1137 otoscopic images, comprising 987 images of normal cases and 150 images of cases of acute OM and OM with effusion, were used as the dataset for image classification. Two convolutional neural network models, trained using our dataset, were used for raw image segmentation and OM classification. RESULTS: The tele-otoscope delivered images with a resolution of 1280 × 720 pixels. Our tele-otoscope effectively differentiated OM from normal images, achieving a classification accuracy rate of up to 94% (sensitivity, 80%; specificity, 96%). CONCLUSION: Our study demonstrated that the developed tele-otoscope has acceptable accuracy in diagnosing OM. This system can assist health care professionals in early detection and continuous remote monitoring, thus mitigating the consequences of OM.

Use of tropical maize for bioethanol production.

Tropical maize is an alternative energy crop being considered as a feedstock for bioethanol production in the North Central and Midwest United States. Tropical maize is advantageous because it produces large amounts of soluble sugars in its stalks, creates a large amount of biomass, and requires lower inputs (e.g. nitrogen) than grain corn. Soluble sugars, including sucrose, glucose and fructose were extracted by pressing the stalks at dough stage (R4). The initial extracted syrup fermented faster than the control culture grown on a yeast extract/phosphate/sucrose medium. The syrup was subsequently concentrated 1.25-2.25 times, supplemented with urea, and fermented using Saccharomyces cerevisiae for up to 96 h. The final ethanol concentrations obtained were 8.1 % (v/v) to 15.6 % (v/v), equivalent to 90.3-92.2 % of the theoretical yields. However, fermentation productivity decreased with sugar concentration, suggesting that the yeast might be osmotically stressed at the increased sugar concentrations. These results provide in-depth information for utilizing tropical maize syrup for bioethanol production that will help in tropical maize breeding and development for use as another feedstock for the biofuel industry.

Differences in fine arabinoxylan structures govern microbial selection and competition among human gut microbiota.

Dietary fibers are known to modulate microbiome composition, but it is unclear to what extent minor fiber structural differences impact community assembly, microbial division of labor, and organismal metabolic responses. To test the hypothesis that fine linkage variations afford different ecological niches for distinct communities and metabolism, we employed a 7-day in vitro sequential batch fecal fermentation with four fecal inocula and measured responses using an integrated multi-omics approach. Two sorghum arabinoxylans (SAXs) were fermented, with one (RSAX) having slightly more complex branch linkages than the other (WSAX). Although there were minor glycoysl linkage differences, consortia on RSAX retained much higher species diversity (42 members) than on WSAX (18-23 members) with distinct species-level genomes and metabolic outcomes (e.g., higher short chain fatty acid production from RSAX and more lactic acid produced from WSAX). The major SAX-selected members were from genera of Bacteroides and Bifidobacterium and family Lachnospiraceae. Carbohydrate active enzyme (CAZyme) genes in metagenomes revealed broad AX-related hydrolytic potentials among key members; however, CAZyme genes enriched in different consortia displayed various catabolic domain fusions with diverse accessory motifs that differ among the two SAX types. These results suggest that fine polysaccharide structure exerts deterministic selection effect for distinct fermenting consortia.

How Many Nodes to Take? Lymph Node Ratio Below 1/3 Reduces Papillary Thyroid Cancer Nodal Recurrence.

INTRODUCTION: Papillary thyroid carcinoma (PTC) accounts for the majority of thyroid malignancies; risk of PTC recurrence over a 30-year period is approximately 30%, of which 70% occur as nodal metastases. Patients with nodal disease who are treated with therapeutic dissection are at higher risk for recurrence, but optimal nodal yield has not been defined. We aim to determine variables predictive of nodal recurrence of PTC within the first 5 years of surgery, with a focus on lymph node ratio (LNR), to inform clinical decision-making. METHODS: Retrospective chart review identified 41 patients with nodal recurrence of PTC and 284 without nodal recurrence following thyroid surgery from 2000 to 2015. Cohorts were compared with regards to clinical history, surgical findings, and tumor characteristics. RESULTS: The fraction of the patients who underwent therapeutic central or lateral lymph node dissection was significantly higher in the nodal recurrence cohort. Maximum tumor size, presence of extrathyroidal extension, largest lymph node focus, LNR, postoperative thyroglobulin level, and administration of postoperative radioactive iodine were significantly increased in the PTC nodal recurrence group. LNR greater than 0.3 held the highest level of significance as a binary cutoff and captured the larger proportion of patients in the nodal recurrence cohort (68.3%). CONCLUSION: This study demonstrates characteristics to help assess risk of nodal recurrence of PTC and suggests LNR of lower than 0.3 is optimal to reduce risk of recurrence. The next steps include cohort studies to validate findings and weight variable analysis to optimize the extent of surgical therapeutic dissection. LEVEL OF EVIDENCE: 4 Laryngoscope, 132:1883-1887, 2022.

Production of bimodal molecular weight levan by a Lactobacillus reuteri isolate from fish gut.

An exopolysaccharide (EPS) synthesizing potentially probiotic Gram-positive bacterial strain was isolated from fish (Tor putitora) gut, and its EPS was structurally characterized. The isolate, designated as FW2, was identified as Lactobacillus reuteri through 16S rRNA gene sequencing and phylogenetic analysis. This isolate produces fructan-type EPS using sucrose as a substrate. Based on 13C-NMR spectroscopy, methylation analysis and monosaccharide composition, the EPS was identified as a linear levan polymer with fructose as main constituent linked via ß(2 → 6) linkages. Based on molecular weight (MW) distribution, two groups of levan were found to be produced by the isolate FW2: one with high MW (4.6 × 106 Da) and the other having much lower MW (1.2 × 104 Da). The isolate yielded about 14 g/L levan under optimized culturing parameters including aeration conditions, pH, temperature and substrate concentration. The obtained bimodal molecular weight linear levan is the first of its type to be synthesized by a L. reuteri isolate from fish gut. Bimodal molecular weight prebiotic levan together with the probiotic potential of the producing strain would provide a new promising synbiotic combination for use in aqua culture.

Absence of the transcriptional repressor Blimp-1 in hematopoietic lineages reveals its role in dendritic cell homeostatic development and function.

Dendritic cells (DCs) are important for the initiation and regulation of immune responses. In this study, we demonstrate that DC homeostatic development in peripheral lymphoid organs is negatively regulated by the transcriptional repressor, Blimp-1, which is critical for regulation of plasma cell differentiation and T cell homeostasis and function. Deletion of Prdm1, the gene encoding Blimp-1, in mouse hematopoietic lineages resulted in an increase in the steady-state number of conventional DCs (cDCs). Specifically, Prdm1 deletion increased immediate CD8(-) cDC precursors in peripheral lymphoid organs, causing selective expansion of the CD8(-) cDC population. Upon stimulus-induced maturation, Blimp-1 was up-regulated in bone marrow-derived DCs via the p38 MAPK and NF-kappaB pathways. Notably, Blimp-1-deficient DCs matured poorly upon stimulation in vitro and in vivo. Blimp-1 binds to the proinflammatory cytokine/chemokine genes, Il-6 and Ccl2, and negatively regulates their expression. Collectively, our findings reveal two new roles for Blimp-1: negative regulation of a select subset of cDCs during homeostatic development, and enhancement of DC maturation.

Controlling autohydrolysis conditions to produce xylan-derived fibers that modulate gut microbiota responses and metabolic outputs.

Autohydrolysis is used for producing xylan-derived oligosaccharides from lignocellulosic biomass. Although numerous studies report optimized autohydrolysis conditions for various plants, few of these studies correlate process parameters with the resulting structural properties to their impact on intestinal bacterial communities. Thus, to further clarify these relationships, beechwood xylan (BWX)-derived substrates, processed under five conditions, were fermented in vitro by human gut microbiota. Autohydrolysis reduced the mean molecular size and substitutions of BWX. Distinct fermentation kinetics were observed with differing processing of BWX substrates, which correlated with impacts on community species evenness. The relative abundances of Bacteroides, Fusicatenibacter, Bifidobacterium, and Megasphaera within the fermentations varied with processing conditions. While the total short-chain fatty acid concentrations were the same among the treatments, processing conditions varied the extent of propionate and butyrate generation. Autolysis parameters may be an important tool for optimizing beneficial effects of xylan-derived fibers on human gut microbiota structure and function.

Fine Carbohydrate Structure of Dietary Resistant Glucans Governs the Structure and Function of Human Gut Microbiota.

Increased dietary fiber consumption has been shown to increase human gut microbial diversity, but the mechanisms driving this effect remain unclear. One possible explanation is that microbes are able to divide metabolic labor in consumption of complex carbohydrates, which are composed of diverse glycosidic linkages that require specific cognate enzymes for degradation. However, as naturally derived fibers vary in both sugar composition and linkage structure, it is challenging to separate out the impact of each of these variables. We hypothesized that fine differences in carbohydrate linkage structure would govern microbial community structure and function independently of variation in glycosyl residue composition. To test this hypothesis, we fermented commercially available soluble resistant glucans, which are uniformly composed of glucose linked in different structural arrangements, in vitro with fecal inocula from each of three individuals. We measured metabolic outputs (pH, gas, and short-chain fatty acid production) and community structure via 16S rRNA amplicon sequencing. We determined that community metabolic outputs from identical glucans were highly individual, emerging from divergent initial microbiome structures. However, specific operational taxonomic units (OTUs) responded similarly in growth responses across individuals' microbiota, though in context-dependent ways; these data suggested that certain taxa were more efficient in competing for some structures than others. Together, these data support the hypothesis that variation in linkage structure, independent of sugar composition, governs compositional and functional responses of microbiota.

Structurally complex carbohydrates maintain diversity in gut-derived microbial consortia under high dilution pressure.

Dietary fibers are major substrates for maintaining and shaping gut microbiota, but the structural specificity of these fibers for the diversity, structure and function of gut microbiota are poorly understood. Here, we employed an in vitro sequential batch fecal culture approach to address two ecological questions: (i) whether the chemical complexity of a carbohydrate influences its ability to maintain microbial diversity against high dilution pressure (ii) whether substrate structuring or obligate microbe-microbe metabolic interactions (e.g. exchange of amino acids or vitamins) exert more influence on maintained diversity. Sorghum arabinoxylan (SAX, a complex polysaccharide), inulin (a low-complexity oligosaccharide) and their corresponding monosaccharide controls were selected as model carbohydrates. Our results demonstrate that complex carbohydrates stably sustain diverse microbial consortia. Furthermore, other metabolic interactions were less influential in structuring microbial consortia consuming SAX than inulin. Finally, very similar final consortia were enriched on SAX from the same individual's fecal microbiota one month later, suggesting that polysaccharide structure is more influential than stochastic alterations in microbiome composition in governing the outcomes of sequential batch cultivation experiments. These data suggest that carbohydrate structural complexity affords independent niches that structure fermenting microbial consortia, whereas other metabolic interactions govern the composition of communities fermenting simpler carbohydrates.

Determining the critical effective temperature and heat dispersal pattern in monopolar radiofrequency ablation using temperature-time integration.

The radiofrequency ablation (RFA) lesion size is posited to be disproportionate to the total delivered energy, and temperature-time integration (TTI) may have a more critical effect on lesion size. The present study aimed to evaluate this hypothesis by determining the temperature threshold and temperature distribution over tissues during the RFA lesioning process. Using an ex vivo chicken tissue model and an in vivo rabbit model with RFA applied for 2 min under various target temperature settings, the resultant lesions were evaluated histologically using Masson's trichrome stain. The temperature distribution over the tissue during the RFA lesioning process was also determined using a VT02 Visual IR Thermometer. It was revealed that the thermal injury threshold for RFA in the chicken tissues was ~65°C, but that it ranged from 55-65°C in mammals. Using infra-red thermal imaging, the temperature gradient (from the center to the periphery) during the RFA lesioning process demonstrated a uniform heat diffusion pattern. This data supports the proposed hypothesis that TTI is a critical parameter in determining RFA lesion size and can be applied clinically using the following equation: [Target temperature - 55 (°C)] × time (sec) is proportional to RFA lesion size.

Miscanthus×giganteus xylooligosaccharides: Purification and fermentation.

A procedure was developed to recover xylooligosaccharides (XOS) from Miscanthus×giganteus (M×G) hydrolyzate. M×G hydrolyzate was prepared using autohydrolysis, and XOS rich fractions were acquired using activated carbon adsorption and stepwise ethanol elution. The combined XOS fractions were purified using a series of ion exchange resin treatments. The end product, M×G XOS, had 89.1% (w/w) total substituted oligosaccharides (TSOS) composed of arabinose, glucose, xylose and acetyl group. Bifidobacterium adolescentis and Bifidobacterium catenulatum (health promoting bacteria) were cultured in vitro on M×G XOS and a commercial XOS source, which was used as a comparison. B. adolescentis grew to a higher cell density than B. catenulatum in both XOS cultures. Total xylose consumption for B. adolescentis was 84.1 and 84.8%, respectively for M×G and commercial XOS cultures; and for B. catenulatum was 76.6 and 73.6%, respectively. The xylobiose (X2), xylotriose (X3) and xylotetraose (X4) were almost utilized for both strains. Acetic and lactic acids were the major fermentation products of the XOS cultures.

In Vitro Fermentation of Xylooligosaccharides Produced from Miscanthus × giganteus by Human Fecal Microbiota.

Purified xylooligosaccharides from Miscanthus × giganteus (M×G XOS) were used in an in vitro fermentation experiment inoculated with human fecal microbiota. A commercial XOS product and pectin were used as controls. Decreases in pH by 2.3, 2.4, and 2.0 units and production of short-chain fatty acids (SCFA; acetic acid, 7764.2, 6664.1, and 6387.9 µmol/g; propionic acid, 1006.7, 1089.5, and 661.5 µmol/g; and butyric acid, 955.5, 1252.9, and 917.7 µmol/g) were observed in M×G XOS, commercial XOS, and pectin medium after 12 h of fermentation, respectively. Titers of Bifidobacterium spp., Lactobacillus spp., and Escherichia coli increased when fed all three substrates as monitored by qPCR. There was no significant trend for Clostridium perfringens. During fermentation, M×G XOS was statistically equivalent in performance to the commercial XOS sample as measured by culture acidification and growth of health-promoting bacteria and resulted in the highest SCFA production among the three substrates.

Ethanol production from food waste at high solids content with vacuum recovery technology.

Ethanol production from food wastes does not only solve environmental issues but also provides renewable biofuels. This study investigated the feasibility of producing ethanol from food wastes at high solids content (35%, w/w). A vacuum recovery system was developed and applied to remove ethanol from fermentation broth to reduce yeast ethanol inhibition. A high concentration of ethanol (144 g/L) was produced by the conventional fermentation of food waste without a vacuum recovery system. When the vacuum recovery is applied to the fermentation process, the ethanol concentration in the fermentation broth was controlled below 100 g/L, thus reducing yeast ethanol inhibition. At the end of the conventional fermentation, the residual glucose in the fermentation broth was 5.7 g/L, indicating incomplete utilization of glucose, while the vacuum fermentation allowed for complete utilization of glucose. The ethanol yield for the vacuum fermentation was found to be 358 g/kg of food waste (dry basis), higher than that for the conventional fermentation at 327 g/kg of food waste (dry basis).

Autohydrolysis of Miscanthus x giganteus for the production of xylooligosaccharides (XOS): kinetics, characterization and recovery.

The optima conditions of production and purification of xylooligosaccharides (XOS) from Miscanthus x giganteus (MxG) were investigated. Using autohydrolysis, XOS were produced at 160, 180 and 200°C at 60, 20 and 5min, respectively. XOS yield up to 13.5% (w/w) of initial biomass and 69.2% (w/w) of xylan were achieved. Results from HPAEC-PAD analysis revealed that X1-X9 sugar oligomers were produced. Higher temperature and longer reaction time resulted in lower product molecular weight. The three optimum conditions had similar degrees of polymerization XOS. Using 10% activated carbon (w/v) with ethanol/water elution recovered 47.9% (w/w) of XOS from pretreated liquid phase. The XOS could be fractionated by degree of polymerization according to ethanol concentration in the ethanol/water elution. Most of the XOS were washed out in 30% and 50% ethanol/water (v/v) fractions. Recoveries of 91.8% xylobiose, 86.9% xylotriose, 66.3% xylotetrose, 56.2% xylopentose and 48.9% xylohexaose were observed in XOS.