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Disposal of electrons generated during the fermentation of ingested feed is a fundamental feature of anaerobic microbial gut ecosystems. Here, we focus on the well-studied rumen environment to highlight how electrons are transferred through anaerobic fermentation pathways and how manipulating this electron flow is important to reducing methane emissions from ruminants. Priorities for research that can accelerate understanding in this area are highlighted.

Molecular hydrogen (H2) and formate (HCOO−) are metabolic end products of many primary fermenters in the mammalian gut. Both play a vital role in fermentation where they are electron sinks for individual microbes in an anaerobic environment that lacks external electron acceptors. If H2 and/or formate accumulate within the gut ecosystem, the ability of primary fermenters to regenerate electron carriers may be inhibited and microbial metabolism and growth disrupted. Consequently, H2- and/or formate-consuming microbes such as methanogens and homoacetogens play a key role in maintaining the metabolic efficiency of primary fermenters. There is increasing interest in identifying approaches to manipulate mammalian gut environments for the benefit of the host and the environment. As H2 and formate are important mediators of interspecies interactions, an understanding of their production and utilisation could be a significant entry point for the development of successful interventions. Ruminant methane mitigation approaches are discussed as a model to help understand the fate of H2 and formate in gut systems.

Background Digestive processes in the rumen lead to the release of methyl-compounds, mainly methanol and methylamines, which are used by methyltrophic methanogens to form methane, an important agricultural greenhouse gas. Methylamines are produced from plant phosphatidylcholine degradation, by choline trimethylamine lyase, while methanol comes from demethoxylation of dietary pectins via pectin methylesterase activity. We have screened rumen metagenomic and metatranscriptomic datasets, metagenome assembled genomes, and the Hungate1000 genomes to identify organisms capable of producing methyl-compounds. We also describe the enrichment of pectin-degrading and methane-forming microbes from sheep rumen contents and the analysis of their genomes via metagenomic assembly. Results Screens of metagenomic data using the protein domains of choline trimethylamine lyase (CutC), and activator protein (CutD) found good matches only to Olsenella umbonata and to Caecibacter, while the Hungate1000 genomes and metagenome assembled genomes from the cattle rumen found bacteria within the phyla Actinobacteria, Firmicutes and Proteobacteria. The cutC and cutD genes clustered with genes that encode structural components of bacterial microcompartment proteins. Prevotella was the dominant genus encoding pectin methyl esterases, with smaller numbers of sequences identified from other fibre-degrading rumen bacteria. Some large pectin methyl esterases (> 2100 aa) were found to be encoded in Butyrivibrio genomes. The pectin-utilising, methane-producing consortium was composed of (i) a putative pectin-degrading bacterium (phylum Tenericutes, class Mollicutes), (ii) a galacturonate-using Sphaerochaeta sp. predicted to produce acetate, lactate, and ethanol, and (iii) a methylotrophic methanogen, Methanosphaera sp., with the ability to form methane via a primary ethanol-dependent, hydrogen-independent, methanogenesis pathway. Conclusions The main bacteria that produce methyl-compounds have been identified in ruminants. Their enzymatic activities can now be targeted with the aim of finding ways to reduce the supply of methyl-compound substrates to methanogens, and thereby limit methylotrophic methanogenesis in the rumen.

Farmed ruminants are the largest source of anthropogenic methane emissions globally. The methanogenic archaea responsible for these emissions use molecular hydrogen (H2), produced during bacterial and eukaryotic carbohydrate fermentation, as their primary energy source. In this work, we used comparative genomic, metatranscriptomic and co-culture-based approaches to gain a system-wide understanding of the organisms and pathways responsible for ruminal H2 metabolism. Two-thirds of sequenced rumen bacterial and archaeal genomes encode enzymes that catalyse H2 production or consumption, including 26 distinct hydrogenase subgroups. Metatranscriptomic analysis confirmed that these hydrogenases are differentially expressed in sheep rumen. Electron-bifurcating [FeFe]-hydrogenases from carbohydrate-fermenting Clostridia (e.g., Ruminococcus) accounted for half of all hydrogenase transcripts. Various H2 uptake pathways were also expressed, including methanogenesis (Methanobrevibacter), fumarate and nitrite reduction (Selenomonas), and acetogenesis (Blautia). Whereas methanogenesis-related transcripts predominated in high methane yield sheep, alternative uptake pathways were significantly upregulated in low methane yield sheep. Complementing these findings, we observed significant differential expression and activity of the hydrogenases of the hydrogenogenic cellulose fermenter Ruminococcus albus and the hydrogenotrophic fumarate reducer Wolinella succinogenes in co-culture compared with pure culture. We conclude that H2 metabolism is a more complex and widespread trait among rumen microorganisms than previously recognised. There is evidence that alternative hydrogenotrophs, including acetogenic and respiratory bacteria, can prosper in the rumen and effectively compete with methanogens for H2. These findings may help to inform ongoing strategies to mitigate methane emissions by increasing flux through alternative H2 uptake pathways, including through animal selection, dietary supplementation and methanogenesis inhibitors.

International audience; Pectin is abundant in modern day diets, as it comprises the middle lamellae and one-third of the dry carbohydrate weight of fruit and vegetable cell walls. Currently there is no specialized model organism for studying pectin fermentation in the human colon, as our collective understanding is informed by versatile glycan-degrading bacteria rather than by specialist pectin degraders. Here we show that the genome of Monoglobus pectinilyticus possesses a highly specialized glycobiome for pectin degradation, unique amongst Firmicutes known to be in the human gut. Its genome encodes a simple set of metabolic pathways relevant to pectin sugar utilization, and its predicted glycobiome comprises an unusual distribution of carbohydrate-active enzymes (CAZymes) with numerous extracellular methyl/acetyl esterases and pectate lyases. We predict the M. pectinilyticus degradative process is facilitated by cell-surface S-layer homology (SLH) domain-containing proteins, which proteomics analysis shows are differentially expressed in response to pectin. Some of these abundant cell surface proteins of M. pectinilyticus share unique modular organizations rarely observed in human gut bacteria, featuring pectin-specific CAZyme domains and the cell wall-anchoring SLH motifs. We observed M. pectinilyticus degrades various pectins, RG-I, and galactan to produce polysaccharide degradation products (PDPs) which are presumably shared with other inhabitants of the human gut microbiome (HGM). This strain occupies a new ecological niche for a primary degrader specialized in foraging a habitually consumed plant glycan, thereby enriching our understanding of the diverse community profile of the HGM.

Bioprocess optimization for cell-based therapies is a resource heavy activity. To reduce the associated cost and time, it is advantageous to carry out process development in small volume systems, with the caveat that such systems be predictive for process scaleup. The transport of oxygen from the gas phase into the culture medium, characterized using the volumetric mass transfer coefficient, kLa, has been identified as a critical parameter for predictive process scaleup. In both large- and small-scale bioreactors, kLa is controlled via mixing, with the method employed dependent upon the size of the reactor. However, existing microplate bioreactor platforms, beneficial for their low working volumes and throughput and automation capabilities, struggle to achieve desired kLa for mammalian cell cultures. Here, we describe the development and testing of a 96-well microplate with integrated Redbud Posts to provide mixing and thus enhanced kLa. Mixing characteristics were investigated, with actuating Redbud Posts shown (visually) to increase convective transport while producing enhanced kLa, providing means to mimic macroscale mammalian cell growth conditions at the microscale. Improved cell growth rates with mixing was demonstrated for two cell types, indicating the potential for this technology to play a valuable role in early stage bioprocess development and optimization.

BackgroundMedulloblastoma (MB) is a rare brain tumor occurring more frequently in children in whom research has been primarily focused. Treatment recommendations in adults are mainly based on retrospective data and pediatric experience; however, molecular features and treatment tolerance differ between the 2 age groups. In adults, prognostic tools are suboptimal, late recurrences are typical, and long-term sequelae remain understudied. Treatment has not adapted to molecular classification advances; thus, the survival rate of adult MB has not improved.MethodsIn 2017, the National Cancer Institute (NCI) received support from the Cancer Moonshot℠ to address the challenges and unmet needs of adults with rare central nervous system tumors through NCI-CONNECT, a program that creates partnerships among patients, health care professionals, researchers, and advocacy organizations. On November 25, 2019, NCI-CONNECT convened leading clinicians and scientists in a workshop to review advances in research, share scientific insights, and discuss clinical challenges in adult MB.ResultsWorking groups identified unmet needs in clinical trial design, tissue acquisition and testing, tumor modeling, and measurement of clinical outcomes.ConclusionsParticipants identified opportunities for collaboration; discussed plans to create a working group of clinicians, researchers, and patient advocates; and developed specific action items to expedite progress in adult MB.

While CAR-T therapy is a growing and promising area of cancer research, it is limited by high cost and the difficulty of consistently culturing T-cells to therapeutically relevant concentrations ex-vivo. Cytokines IL-2, IL-7 and IL-15 have been found to stimulate the growth of T cells, however, the optimized combination of these three cytokines for T cell proliferation is unknown. In this study, we designed an integrated experimental and modeling approach to optimize cytokine supplementation for rapid expansion in clinical applications. We assessed the growth data for statistical improvements over no cytokine supplementation and used a systems biology approach to identify genes with the highest magnitude of expression change from control at several time points. Further, we developed a predictive mathematical model to project the growth rate for various cytokine combinations, and investigate genes and reactions regulated by cytokines in activated CD4+ T cells. The most favorable conditions from the T cell growth study and from the predictive model align to include the full range of IL-2 and IL-7 studied, and at lower levels of IL-15 (6 ng/mL or 36 ng/mL). The highest growth rates were observed where either IL-2 or IL-7 was at the highest concentration tested (15 ng/mL for IL-2 and 80 ng/mL for IL-7) while the other was at the lowest (1 ng/mL for IL-2 and 6 ng/mL for IL-7), or where both IL-2 and IL-7 concentrations are moderate-corresponding to condition keys 200, 020, and 110 respectively. This suggests a synergistic interaction of IL-2 and IL-7 with regards to promoting optimal proliferation and survival of the activated CD4+ T cells. Transcriptomic data analysis identified the genes and transcriptional regulators up/down-regulated by each of the cytokines IL-2, IL-7, and IL-15. It was found that the genes with persistent expressing changes were associated with major pathways involved in cell growth and proliferation. In addition to influencing T cell metabolism, the three cytokines were found to regulate specific genes involved in TCR, JAK/STAT, MAPK, AKT and PI3K-AKT signaling. The developed Fuzzy model that can predict the growth rate of activated CD4+ T cells for various combinations of cytokines, along with identified optimal cytokine cocktails and important genes found in transcriptomic data, can pave the way for optimizing activated CD4 T cells by regulating cytokines in the clinical setting.

Bacterial species belonging to the genus Pseudobutyrivibrio are important members of the rumen microbiome contributing to the degradation of complex plant polysaccharides. Pseudobutyrivibrio xylanivorans MA3014 was selected for genome sequencing to examine its ability to breakdown and utilize plant polysaccharides. The complete genome sequence of MA3014 is 3.58 Mb, consists of three replicons (a chromosome, chromid, and plasmid), has an overall G + C content of 39.6%, and encodes 3,265 putative protein-coding genes (CDS). Comparative pan-genomic analysis of all cultivated and currently available P. xylanivorans genomes has revealed a strong correlation of orthologous genes within this rumen bacterial species. MA3014 is metabolically versatile and capable of growing on a range of simple mono- or oligosaccharides derived from complex plant polysaccharides such as pectins, mannans, starch, and hemicelluloses, with lactate, butyrate, and formate as the principal fermentation end products. The genes encoding these metabolic pathways have been identified and MA3014 is predicted to encode an extensive range of Carbohydrate-Active enZYmes with 78 glycoside hydrolases, 13 carbohydrate esterases, and 54 glycosyl transferases, suggesting an important role in solubilization of plant matter in the rumen.

Ruminants are essential for maintaining the global population and managing greenhouse gas emissions. In the rumen, bacterial species belonging to the genera rumen Butyrivibrio and Pseudobutyrivibrio constitute the core bacterial rumen microbiome and are important degraders of plant-derived complex polysaccharides. Pseudobutyrivibrio xylanivorans MA3014 was selected for genome sequencing in order to examine its ability to breakdown and utilize plant polysaccharides. The complete genome sequence of MA3014 is 3.58 Mb, consists of three replicons (a chromosome, chromid and plasmid), has an overall G+C content of 39.6% and encodes 3,265 putative protein-coding genes (PCGs). Comparative pan-genomics of all cultivated and currently available P. xylanivorans genomes has revealed highly open genomes and a strong correlation of orthologous genes within this species of rumen bacteria. MA3014 is metabolically versatile and capable of utilizing a range of simple mono-or oligosaccharides to complex plant polysaccharides such as pectins, mannans, starch and hemicelluloses for growth, with lactate, butyrate and formate as the principal fermentation end-products. The genes encoding these metabolic pathways have been identified and MA3014 is predicted to encode an extensive repertoire of Carbohydrate-Active enZYmes (CAZymes) with 80 Glycoside Hydrolases (GHs), 28 Carbohydrate Esterases (CEs) and 51 Glycosyl Transferases (GTs), that suggest its role as an initiator of primary solubilization of plant matter in the rumen.

Immunotherapy, activation of the immune system to target tumor cells, represents a paradigm shift in the treatment of cancer. Immune checkpoint therapies, which target immunomodulatory molecules expressed on T-lymphocytes, have demonstrated improved survival in a variety of malignancies. However, benefit in glioblastoma, the most common and devastating malignant brain tumor, remains to be seen. With several recent clinical trials failing to show efficacy of immunotherapy, concerns have been raised regarding the impact of glucocorticoid use in this patient population that may impair the ability for immune checkpoint inhibitors to affect a response. For this article we examined the mechanism by which immune checkpoint inhibitors activate, and glucocorticoids impair, T-lymphocyte function. In this context, we review the clinical data of immune checkpoint inhibitors in glioblastoma as well as the impact glucocorticoids have on immune checkpoint inhibitor efficacy. Finally, we highlight key questions that remain in the field, and the potential benefit of further research for central nervous system tumors. More information on the extent, character and duration of glucocorticoids on patients treated with PD-(L)1 will better inform both clinical management and novel therapeutic development of immunotherapy in patients with CNS malignancies.

It is known that process scaling-up has always been a challenge in biopharmaceutical and food industry. In recent years, newly emerging microscale bioreactors like Ambr15 and Ambr250 have attracted significant attention for that they can provide high throughput and accelerate upstream process development. In this work, we developed the first multiphase Computational Fluid Dynamics (CFD) model for an in-depth characterization of Ambr bioreactor systems. A number of advanced computational methods, including Reynolds stress turbulence model, population balance model, multiple reference frame (MRF), sliding mesh (SM) and user defined functions (UDFs), were integrated for the first time to systematically study the gas-liquid mixing in Ambr250 bioreactor. We provided detailed comparison between MRF and SM method, demonstrated the limitation of MRF for predicting bubble distribution in asymmetric reactors. Characteristics of hydrodynamics, mass transfer, turbulent dissipation and bubble size distribution were predicted from our CFD models and validated by existing experimental data for a variety of operating conditions for both the Ambr15 and Ambr250 bioreactors. The predicted kLa value ranges are 0.18–7.90 h-1 and 2.15–11.52 h-1 for Ambr250 and Ambr15, respectively. This work thus provides a superior framework for the computational modeling of microscale stirred bioreactors.

Plant polysaccharide breakdown by microbes in the rumen is fundamental to digestion in ruminant livestock. Bacterial species belonging to the rumen genera Butyrivibrio and Pseudobutyrivibrio are important degraders and utilizers of lignocellulosic plant material. These bacteria degrade polysaccharides and ferment the released monosaccharides to yield short-chain fatty acids that are used by the ruminant for growth and the production of meat, milk, and fiber products. Although rumen Butyrivibrio and Pseudobutyrivibrio species are regarded as common rumen inhabitants, their polysaccharide-degrading and carbohydrate-utilizing enzymes are not well understood. In this study, we analyzed the genomes of 40 Butyrivibrio and 6 Pseudobutyrivibrio strains isolated from the plant-adherent fraction of New Zealand dairy cows to explore the polysaccharide-degrading potential of these important rumen bacteria. Comparative genome analyses combined with phylogenetic analysis of their 16S rRNA genes and short-chain fatty acid production patterns provide insight into the genomic diversity and physiology of these bacteria and divide Butyrivibrio into 3 species clusters. Rumen Butyrivibrio bacteria were found to encode a large and diverse spectrum of degradative carbohydrate-active enzymes (CAZymes) and binding proteins. In total, 4,421 glycoside hydrolases (GHs), 1,283 carbohydrate esterases (CEs), 110 polysaccharide lyases (PLs), 3,605 glycosyltransferases (GTs), and 1,706 carbohydrate-binding protein modules (CBM) with predicted activities involved in the depolymerization and transport of the insoluble plant polysaccharides were identified. Butyrivibrio genomes had similar patterns of CAZyme families but varied greatly in the number of genes within each category in the Carbohydrate-Active Enzymes database (CAZy), suggesting some level of functional redundancy. These results suggest that rumen Butyrivibrio species occupy similar niches but apply different degradation strategies to be able to coexist in the rumen.IMPORTANCE Feeding a global population of 8 billion people and climate change are the primary challenges facing agriculture today. Ruminant livestock are important food-producing animals, and maximizing their productivity requires an understanding of their digestive systems and the roles played by rumen microbes in plant polysaccharide degradation. Members of the genera Butyrivibrio and Pseudobutyrivibrio are a phylogenetically diverse group of bacteria and are commonly found in the rumen, where they are a substantial source of polysaccharide-degrading enzymes for the depolymerization of lignocellulosic material. Our findings have highlighted the immense enzymatic machinery of Butyrivibrio and Pseudobutyrivibrio species for the degradation of plant fiber, suggesting that these bacteria occupy similar niches but apply different degradation strategies in order to coexist in the competitive rumen environment.

Membrane chromatography is an emerging bioprocessing technology. This work aimed to determine the cause of pressure rise during the loading phase of a large-scale viral recovery process, through small-scale experimentation. Comparable pressure rise was observed at both small and large scales, using Sartobind® Q anion exchange membranes. By varying low rate and feed concentrations of virus and host cell DNA, a linear relation between Viral Dynamic Dynamic Binding Capacity (DBC) and pressure rise was observed when DNA levels exceeded 1 × 10−8 g/ml. A mathematical model, assuming an uncompetitive interaction between the two species in a representative pore, was developed to predict breakthrough from which DBC and then pressure rise was determined. The results matched experimental data quite well at low to moderate DBC values (ie

The ambr bioreactors are single-use microbioreactors for cell line development and process optimization. With operating conditions for large-scale biopharmaceutical production properly scaled down, microbioreactors such as the ambr15™ can potentially be used to predict the effect of process changes such as modified media or different cell lines. While there have been some recent studies evaluating the ambr15™ technology as a scale-down model for fed-batch operations, little has been reported for semi-continuous or continuous operation. Gassing rates and dilution rates in the ambr15™ were varied in this study to attempt to replicate performance of a perfusion process at the 5 L scale. At both scales, changes to metabolite production and consumption, and cell growth rate and therapeutic protein production were measured. Conditions were identified in the ambr15™ bioreactor that produced metabolic shifts and specific metabolic and protein production rates that are characteristic of the corresponding 5 L perfusion process. A dynamic flux balance (DFB) model was employed to understand and predict the metabolic changes observed. The DFB model predicted trends observed experimentally, including lower specific glucose consumption and a switch from lactate production to consumption when dissolved CO2 was maintained at higher levels in the broth. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 34:420-431, 2018.

The amount and ratio of organic acids from carbohydrate fermentation by the gut microbiota is dependent upon both the ability of members of the microbiota to exploit undigested dietary carbohydrate and the type of organic acids they have the capability to generate. Additionally, some bacteria feed upon carbohydrate degradation products, as well as organic acids produced by other bacteria, further affecting the overall organic acid profile. Experimentally unentangling these complex trophic food webs is challenging. Here we demonstrate a mathematical model that calculates the organic acid profile resulting from an in vitro human faecal microbial fermentation of kiwifruit cell wall polysaccharides, parameterised with organic acid and microbial abundance data. The model was based on our hypothesis that in conditions of carbohydrate excess, changes in the abundance of members of the microbiota will be reflected in an equivalent change in those organic acids they have the capability to produce. We used microbial abundance data to calculate changes in organic acids over time, and the resulting data was shown to recapitulate the measured organic acid profiles. We also demonstrate here the successful mathematical simulation of the metabolic cross-feeding which occurs upon simulated digestion and fermentation of a fresh whole fruit carbohydrate. We assign the relative propensity of various members of a complex microbial community to generate selected acids within the organic acid profiles. We anticipate this case study to be a starting point for more sophisticated research and clinical tools to investigate and predict the interactions with food, the microbiota and the host.

Feeding a global population of 8 billion people and climate change are the primary challenges facing agriculture today. Ruminant livestock are important food-producing animals, and maximizing their productivity requires an understanding of their digestive systems and the roles played by rumen microbes in plant polysaccharide degradation. Members of the genera Butyrivibrio and Pseudobutyrivibrio are a phylogenetically diverse group of bacteria and are commonly found in the rumen, where they are a substantial source of polysaccharide-degrading enzymes for the depolymerization of lignocellulosic material. Our findings have highlighted the immense enzymatic machinery of Butyrivibrio and Pseudobutyrivibrio species for the degradation of plant fiber, suggesting that these bacteria occupy similar niches but apply different degradation strategies in order to coexist in the competitive rumen environment., Plant polysaccharide breakdown by microbes in the rumen is fundamental to digestion in ruminant livestock. Bacterial species belonging to the rumen genera Butyrivibrio and Pseudobutyrivibrio are important degraders and utilizers of lignocellulosic plant material. These bacteria degrade polysaccharides and ferment the released monosaccharides to yield short-chain fatty acids that are used by the ruminant for growth and the production of meat, milk, and fiber products. Although rumen Butyrivibrio and Pseudobutyrivibrio species are regarded as common rumen inhabitants, their polysaccharide-degrading and carbohydrate-utilizing enzymes are not well understood. In this study, we analyzed the genomes of 40 Butyrivibrio and 6 Pseudobutyrivibrio strains isolated from the plant-adherent fraction of New Zealand dairy cows to explore the polysaccharide-degrading potential of these important rumen bacteria. Comparative genome analyses combined with phylogenetic analysis of their 16S rRNA genes and short-chain fatty acid production patterns provide insight into the genomic diversity and physiology of these bacteria and divide Butyrivibrio into 3 species clusters. Rumen Butyrivibrio bacteria were found to encode a large and diverse spectrum of degradative carbohydrate-active enzymes (CAZymes) and binding proteins. In total, 4,421 glycoside hydrolases (GHs), 1,283 carbohydrate esterases (CEs), 110 polysaccharide lyases (PLs), 3,605 glycosyltransferases (GTs), and 1,706 carbohydrate-binding protein modules (CBM) with predicted activities involved in the depolymerization and transport of the insoluble plant polysaccharides were identified. Butyrivibrio genomes had similar patterns of CAZyme families but varied greatly in the number of genes within each category in the Carbohydrate-Active Enzymes database (CAZy), suggesting some level of functional redundancy. These results suggest that rumen Butyrivibrio species occupy similar niches but apply different degradation strategies to be able to coexist in the rumen. IMPORTANCE Feeding a global population of 8 billion people and climate change are the primary challenges facing agriculture today. Ruminant livestock are important food-producing animals, and maximizing their productivity requires an understanding of their digestive systems and the roles played by rumen microbes in plant polysaccharide degradation. Members of the genera Butyrivibrio and Pseudobutyrivibrio are a phylogenetically diverse group of bacteria and are commonly found in the rumen, where they are a substantial source of polysaccharide-degrading enzymes for the depolymerization of lignocellulosic material. Our findings have highlighted the immense enzymatic machinery of Butyrivibrio and Pseudobutyrivibrio species for the degradation of plant fiber, suggesting that these bacteria occupy similar niches but apply different degradation strategies in order to coexist in the competitive rumen environment.

The taxonomy and associated nomenclature of many taxa of rumen bacteria are poorly defined within databases of 16S rRNA genes. This lack of resolution results in inadequate definition of microbial community structures, with large parts of the community designated as incertae sedis, unclassified, or uncultured within families, orders, or even classes. We have begun resolving these poorly-defined groups of rumen bacteria, based on our desire to name these for use in microbial community profiling. We used the previously-reported global rumen census (GRC) dataset consisting of >4.5 million partial bacterial 16S rRNA gene sequences amplified from 684 rumen samples and representing a wide range of animal hosts and diets. Representative sequences from the 8,985 largest operational units (groups of sequence sharing >97% sequence similarity, and covering 97.8% of all sequences in the GRC dataset) were used to identify 241 pre-defined clusters (mainly at genus or family level) of abundant rumen bacteria in the ARB SILVA 119 framework. A total of 99 of these clusters (containing 63.8% of all GRC sequences) had no unique or had inadequate taxonomic identifiers, and each was given a unique nomenclature. We assessed this improved framework by comparing taxonomic assignments of bacterial 16S rRNA gene sequence data in the GRC dataset with those made using the original SILVA 119 framework, and three other frameworks. The two SILVA frameworks performed best at assigning sequences to genus-level taxa. The SILVA 119 framework allowed 55.4% of the sequence data to be assigned to 751 uniquely identifiable genus-level groups. The improved framework increased this to 87.1% of all sequences being assigned to one of 871 uniquely identifiable genus-level groups. The new designations were included in the SILVA 123 release (https://www.arb-silva.de/documentation/release-123/) and will be perpetuated in future releases.

Feeding a future global population of 9 billion people and climate change are the primary challenges facing agriculture today. Ruminant livestock are important food-producing animals, and maximizing their productivity requires an understanding of their digestive systems and the roles played by rumen microbes in plant polysaccharide degradation. Butyrivibrio species are a phylogenetically diverse group of bacteria and are commonly found in the rumen, where they are a substantial source of polysaccharide-degrading enzymes for the depolymerization of lignocellulosic material. Our findings suggest that closely related species of Butyrivibrio have developed unique strategies for the degradation of plant fiber and the subsequent assimilation of carbohydrates in order to coexist in the competitive rumen environment. The identification of genes expressed during these competitive interactions gives further insight into the enzymatic machinery used by these bacteria as they degrade the xylan and pectin components of plant fiber., Rumen bacterial species belonging to the genus Butyrivibrio are important degraders of plant polysaccharides, particularly hemicelluloses (arabinoxylans) and pectin. Currently, four species are recognized; they have very similar substrate utilization profiles, but little is known about how these microorganisms are able to coexist in the rumen. To investigate this question, Butyrivibrio hungatei MB2003 and Butyrivibrio proteoclasticus B316T were grown alone or in coculture on xylan or pectin, and their growth, release of sugars, fermentation end products, and transcriptomes were examined. In monocultures, B316T was able to grow well on xylan and pectin, while MB2003 was unable to utilize either of these insoluble substrates to support significant growth. Cocultures of B316T grown with MB2003 revealed that MB2003 showed growth almost equivalent to that of B316T when either xylan or pectin was supplied as the substrate. The effect of coculture on the transcriptomes of B316T and MB2003 was assessed; B316T transcription was largely unaffected by the presence of MB2003, but MB2003 expressed a wide range of genes encoding proteins for carbohydrate degradation, central metabolism, oligosaccharide transport, and substrate assimilation, in order to compete with B316T for the released sugars. These results suggest that B316T has a role as an initiator of primary solubilization of xylan and pectin, while MB2003 competes effectively for the released soluble sugars to enable its growth and maintenance in the rumen. IMPORTANCE Feeding a future global population of 9 billion people and climate change are the primary challenges facing agriculture today. Ruminant livestock are important food-producing animals, and maximizing their productivity requires an understanding of their digestive systems and the roles played by rumen microbes in plant polysaccharide degradation. Butyrivibrio species are a phylogenetically diverse group of bacteria and are commonly found in the rumen, where they are a substantial source of polysaccharide-degrading enzymes for the depolymerization of lignocellulosic material. Our findings suggest that closely related species of Butyrivibrio have developed unique strategies for the degradation of plant fiber and the subsequent assimilation of carbohydrates in order to coexist in the competitive rumen environment. The identification of genes expressed during these competitive interactions gives further insight into the enzymatic machinery used by these bacteria as they degrade the xylan and pectin components of plant fiber.

Farmed ruminants are the largest source of anthropogenic methane emissions globally. The methanogenic archaea responsible for these emissions use molecular hydrogen (H2), produced during bacterial and eukaryotic carbohydrate fermentation, as their primary energy source. In this work, we used comparative genomic, metatranscriptomic, and co-culture-based approaches to gain a system-wide understanding of the organisms and pathways responsible for ruminal H2metabolism. Two thirds of sequenced rumen bacterial and archaeal genomes encode enzymes that catalyze H2production or consumption, including 26 distinct hydrogenase subgroups. Metatranscriptomic analysis confirmed that these hydrogenases are differentially expressed in sheep rumen. Electron-bifurcating [FeFe]-hydrogenases from carbohydrate-fermenting Clostridia (e.g.Ruminococcus) accounted for half of all hydrogenase transcripts. Various H2uptake pathways were also expressed, including methanogenesis (Methanobrevibacter), fumarate reduction and nitrate ammonification (Selenomonas), and acetogenesis (Blautia). Whereas methanogenesis predominated in high methane yield sheep, alternative uptake pathways were significantly upregulated in low methane yield sheep. Complementing these findings, we observed significant differential expression and activity of the hydrogenases of the hydrogenogenic cellulose fermenterRuminococcus albusand the hydrogenotrophic fumarate reducerWolinella succinogenesin co-culture compared to pure culture. We conclude that H2metabolism is a more complex and widespread trait among rumen microorganisms than previously recognized. There is evidence that alternative hydrogenotrophs, including acetogens and selenomonads, can prosper in the rumen and effectively compete with methanogens for H2in low methane yield ruminants. Strategies to increase flux through alternative H2uptake pathways, including animal selection, dietary supplementation, and methanogenesis inhibitors, may lead to sustained methane mitigation.

A combination of biochemical and thermochemical process to produce sugars and bio-oil by using switchgrass which was grown in lead contaminated soil was studied. Four different process routes which involve fast pyrolysis, enzymatic hydrolysis, acid hydrolysis, and their combinations were investigated. The impact of lead content in the switchgrass feedstock used for the production of sugars and bio-oil along with the recovery of lead through the above mentioned routes were evaluated. The yields of sugars from acid hydrolysis and enzymatic hydrolysis did not seem to be affected by the lead contained in the switchgrass. The chemical product distribution in bio-oil produced from pyrolysis of lead-containing switchgrass and regular switchgrass showed slight differences. The highest overall yields of the liquid products were obtained from the fourth route which utilized all the three process steps, resulting in the lowest yield of bio-char byproduct which had the lowest content lead (2 mg/kg).

Rumen bacterial species belonging to the generaButyrivibrioare important degraders of plant polysaccharides, particularly hemicelluloses (arabinoxylans) and pectin. Currently, four distinct species are recognized which have very similar substrate utilization profiles, but little is known about how these microorganisms are able to co-exist in the rumen. To investigate this question,Butyrivibrio hungatei(MB2003) andButyrivibrio proteoclasticus(B316T) were grown alone or in co-culture on the insoluble substrates, xylan or pectin, and their growth, release of sugars, fermentation end products and transcriptomes were examined. In single cultures, B316Twas able to degrade and grow well on xylan and pectin, whileB. hungateiMB2003 was unable to utilize either of these insoluble substrates to support significant growth. Co-cultures of B316Tgrown with MB2003 revealed that MB2003 showed almost equivalent growth to B316Twhen either xylan or pectin were supplied as substrates. The effect of co-culture on the transcriptomes of B316Tand MB2003 was very marked; B316Ttranscription was largely unaffected by the presence MB2003, but MB2003 expressed a wide range of genes encoding carbohydrate degradation/metabolism and oligosaccharide transport/assimilation in order to compete with B316Tfor the released sugars. These results suggest that B316Thas a role as an initiator of the primary solubilization of xylan and pectin, while MB2003 competes effectively as a scavenger for the released soluble sugars to enable its growth and maintenance in the rumen.IMPORTANCEFeeding a global population of nine billion people and climate change are the primary challenges facing agriculture today. Determining the roles of rumen microbes involved in plant polysaccharide breakdown is fundamental to understanding digestion and maximizing productivity of ruminant livestock.Butyrivibrioare abundant rumen bacteria and are a substantial source of polysaccharide-degrading enzymes with biotechnological applications for the depolymerization of lignocellulosic material. Our findings suggest that closely related species ofButyrivibriohave developed unique strategies for the degradation of plant fibre and the subsequent assimilation of carbohydrates in order to coexist in the competitive rumen environment. The identification of genes related to their enzymatic machinery by which these bacteria work in concert to degrade these different forms of polysaccharides contributes to our understanding of carbon flow in the rumen.

Starches resistant to mammalian digestion are present in foods and pass to the large bowel, where they may be degraded and fermented by the microbiota. Increases in relative abundances of bifidobacteria (blooms) have been reported in rats whose diet was supplemented with Hi-Maize resistant starch. We determined that the bifidobacterial species present in the rat cecum under these circumstances mostly belonged to Bifidobacterium animalis However, cultures of B. animalis isolated from the rats failed to degrade Hi-Maize starch to any extent. In contrast, Bifidobacterium pseudolongum also detected in the rat microbiota had high starch-degrading ability. Transcriptional comparisons showed increased expression of a type 1 pullulanase, alpha-amylase, and glycogen debranching enzyme by B. pseudolongum when cultured in medium containing Hi-Maize starch. Maltose was released into the culture medium, and B. animalis cultures had shorter doubling times in maltose medium than did B. pseudolongum Thus, B. pseudolongum, which was present at a consistently low abundance in the microbiota, but which has extensive enzymatic capacity to degrade resistant starch, showed the attributes of a keystone species associated with the bifidobacterial bloom.IMPORTANCE This study addresses the microbiology and function of a natural ecosystem (the rat gut) using DNA-based observations and in vitro experimentation. The microbial community of the large bowel of animals, including humans, has been studied extensively through the use of high-throughput DNA sequencing methods and advanced bioinformatics analysis. These studies reveal the compositions and genetic capacities of microbiotas but not the intricacies of how microbial communities function. Our work, combining DNA sequence analysis and laboratory experiments with cultured strains of bacteria, revealed that the increased abundance of bifidobacteria in the rat gut, induced by feeding indigestible starch, involved a species that cannot itself degrade the starch (Bifidobacterium animalis) but cohabits with a species that can (Bifidobacterium pseudolongum). B. pseudolongum has the characteristics of a keystone species in the community because it had low abundance but high ability to perform a critical function, the hydrolysis of resistant starch.

Immunotherapy as a therapeutic strategy has seized the narrative throughout clinical oncology over the past few years. Once considered a niche treatment for rare cancers, immunotherapy has quickly emerged as the standard of care for many common cancer types. The remarkable rise is largely due to the development of novel checkpoint inhibitors, specifically, antibodies targeting PD-1 and PD-L1. Offering promising efficacy with a favorable toxicity profile, these agents have been approved for use in several malignancies and are under investigation for many more. One of the more appealing features is the chance for meaningful, durable response – uncharacteristic for most cancer therapies. Atezolizumab is a humanized IgG1 monoclonal antibody that targets PD-L1. Atezolizumab has been approved for use in the treatment of advanced non-small cell lung cancer (NSCLC) and bladder cancer and has shown promising activity in several other types of cancer. Here, we provide a product review for atezolizumab.

Butyrivibrio hungatei MB2003 was isolated from the plant-adherent fraction of rumen contents from a pasture-grazed New Zealand dairy cow, and was selected for genome sequencing in order to examine its ability to degrade plant polysaccharides. The genome of MB2003 is 3.39 Mb and consists of four replicons; a chromosome, a secondary chromosome or chromid, a megaplasmid and a small plasmid. The genome has an average G + C content of 39.7%, and encodes 2983 putative protein-coding genes. MB2003 is able to use a variety of monosaccharide substrates for growth, with acetate, butyrate and formate as the principal fermentation end-products, and the genes encoding these metabolic pathways have been identified. MB2003 is predicted to encode an extensive repertoire of CAZymes with 78 GHs, 7 CEs, 1 PL and 78 GTs. MB2003 is unable to grow on xylan or pectin, and its role in the rumen appears to be as a utilizer of monosaccharides, disaccharides and oligosaccharides made available by the degradative activities of other bacterial species. Electronic supplementary material The online version of this article (10.1186/s40793-017-0285-8) contains supplementary material, which is available to authorized users.