Biology subjects

Walter, J.

Publications and source records attributed to Walter, J..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Extensive photophysiological variation in wild barley is linked to environmental origin

O_LIIntraspecific variation between crop wild relatives (CWRs) represents a source of untapped genetic diversity for crop improvement. At the same time, improving photosynthesis in crops has the potential to enhance yield. Thus, exploring variation for photophysiology within CWRs is an important, yet underexplored, research area. C_LIO_LIWe describe a common garden experiment where 320 wild barley accessions were grown across two seasons. A photophysiology phenotyping pipeline was employed to quantify >30 traits within this diversity panel. Population genetics, genome-wide association analyses (GWAS) and deep phenotyping were performed to address local adaptation hypotheses. C_LIO_LIHeritable variation was detected across this photophysiological spectrum, with genotype-by-environment (GxE) interactions being prevalent. Evidence for local adaptation was observed in the form of subpopulation differences, signals of selection, and allele frequency variation associated to markers identified via GWAS. Phenotyping of representative accessions across distinct water availabilities highlighted a role for stomatal conductance (gs) in adaptation to dry environments. C_LIO_LIWe identified substantial variation in key photosynthesis-associated traits in a CWR closely related to barley, an economically important crop species. Our results demonstrate that this variation is partially due to local adaptation, where plasticity in gs appears important for maintaining photosynthesis and biomass accumulation in water restricted conditions. C_LI

plant biology↗

bioRxiv2025

Chloroplast-derived hydrogen peroxide coordinates photosynthesis with stomatal opening

Stomatal pores on plant leaves control the entry point of carbon into the terrestrial biosphere and concomitant release of water vapour into the atmosphere. Stomatal aperture changes need to be coordinated with photosynthesis to balance carbon gain against water loss. Despite this importance, molecular signals underpinning this coordination, and the cell types from which such signals originate, remain poorly understood. Here we show that chloroplast-derived H2O2 coordinates stomatal movements with photosynthesis. Specifically, when H2O2 scavenging is compromised in the cytosolic ascorbate peroxidase1 (apx1) mutant, stomatal opening is increased. This is rescued when photosynthetic H2O2 production derived from plastoquinol is reduced either genetically or pharmacologically. Furthermore, cell-specific complementation of APX1 in the apx1 mutant demonstrated that H2O2 signals from both guard cells and mesophyll cells contribute significantly to stomatal movements. Altogether, our findings identify H2O2 as a key photosynthesis-derived signal underpinning effective coordination between stomatal conductance and photosynthesis, a key prerequisite for terrestrial plant life, and may inspire new strategies to enhance photosynthetic water use efficiency in crop species.

plant biology↗

bioRxiv2025

Epitope Sequence and Modification Fingerprints of Anti-Aβ Antibodies

A hallmark of Alzheimers disease (AD), the most common form of dementia, is the progressive accumulation of amyloid-beta (A{beta}) peptides across distinct brain regions. Anti-A{beta} antibodies (A{beta}-Abs) targeting specific A{beta} variants are essential tools for AD research, diagnostics, and therapy. The monoclonal antibodies Aducanumab, Lecanemab, and Donanemab have recently been approved as the first disease-modifying treatments for early AD, highlighting the clinical importance of their exact binding profiles. In this study, we systematically characterized the binding and modification requirements of 20 A{beta}-Abs, including biosimilars of Aducanumab, Lecanemab, and Donanemab, across monomeric, oligomeric, and aggregated A{beta} forms. Array-based analysis of 20,000 modified A{beta} peptides defined binding epitopes at single-residue resolution and revealed the impact of sequence variation, including familial AD mutations, as well as diverse post-translational modifications (PTMs). Notably, genetic variants such as H6R impaired binding of therapeutic A{beta}-Abs like Aducanumab. Donanemab showed strong preference for pyroglutamate-modified A{beta}pE3-10, while Lecanemab and Aducanumab exhibited aggregation- and sequence-context-dependent binding requirements. Comparison of peptide binding profiles with binding of full-length and aggregated A{beta} via immunoprecipitation-mass spectrometry, capillary immunoassays, Western blotting, and immunohistochemistry on AD brain tissue revealed distinct aggregation-dependent binding behaviours. The valency- and context-dependence of Aducanumab binding, together with its preference for Ser8-phosphorylated A{beta}, supports a dimerization-mediated binding mechanism. For Lecanemab, our data suggest that additional structural contributions beyond the minimal N-terminal epitope are required for binding to aggregated A{beta}, which remain to be fully resolved. Together, this work provides the most comprehensive dataset to date on aggregation-dependent sequence and modification selectivity of A{beta}-Abs. By integrating mutational, PTM, and aggregation contexts in a unified experimental framework, we establish a resource that enables rational selection of antibodies for research and diagnostic applications, and offers mechanistic insights that may inform the design and optimization of future therapeutic antibodies in AD.

biochemistry↗

bioRxiv2025

EndoMAPV1, a Structural Protein Complex Landscape of Human Endosomes

Early/sorting endosomes are dynamic organelles that play key roles in proteome control by triaging plasma membrane proteins for either recycling or degradation in the lysosome1,2,3. These events are coordinated by numerous transiently-associated regulatory complexes and integral membrane components that contribute to organelle identity during endosome maturation4. While a subset of the several hundred protein components and cargoes known to associate with endosomes have been studied at the biochemical and/or structural level, interaction partners and higher order molecular assemblies for many endosomal components remain unknown. Here, we combine cross-linking and native gel mass spectrometry5-8 of purified early endosomes with AlphaFold9,10 and computational analysis to create a systematic human endosomal structural interactome. We present dozens of structural models for endosomal protein pairs and higher order assemblies supported by experimental cross-links from their native subcellular context, suggesting structural mechanisms for previously reported regulatory processes. Using induced neurons, we validate two candidate complexes whose interactions are supported by crosslinks and structural predictions: TMEM230 as a subunit of ATP8/11 lipid flippases11 and TMEM9/9B as subunits of CLCN3/4/5 chloride-proton antiporters12. This resource and its accompanying structural network viewer provide an experimental framework for understanding organellar structural interactomes and large-scale validation of structural predictions.

cell biology↗

bioRxiv2024

Human shape perception spontaneously discovers the biological origin of novel, but natural, stimuli

Humans excel at categorizing objects by shape. This facility involves identifying shape features that objects have in common with other members of their class and relies--at least in part--on semantic/cognitive constructs. For example, plants sprout branches, fish grow fins, shoes are molded to our feet. Can humans parse shapes according to the processes that give shapes their key characteristics, even when such processes are hidden? To answer this, we investigated how humans perceive the shape of cells from the olfactory system of Xenopus laevis tadpoles. These objects are novel to most humans yet occur in nature and cluster into classes following their underlying biological function. We reconstructed 3D cell models through 3D-microscopy and photogrammetry, then conducted psychophysical experiments. Human participants performed two tasks: they arranged 3D-printed cell models by similarity and rated them along eight visual dimensions. Participants were highly consistent in their arrangements and ratings and spontaneously grouped stimuli to reflect the cell classes, unwittingly revealing the underlying processes shaping these forms. Our findings thus demonstrate that human perceptual organization mechanisms spontaneously parse the biological systematicities of never-before-seen, natural shapes. Integrating such human perceptual strategies into automated systems may enhance morphology-based analysis in biology and medicine.

neuroscience↗

bioRxiv2024

TTF2 promotes replisome eviction from stalled forks in mitosis

Cells entering mitosis with incompletely replicated DNA face catastrophic chromosome segregation failure. During interphase, the replisome-associated E3 ubiquitin ligase TRAIP ubiquitylates barriers in front of the fork to allow replisome progression. In mitosis, TRAIP is reprogrammed from a trans-acting to a cis-acting ligase that can ubiquitylate the replisome itself. This enables the processing of unreplicated DNA by promoting replisome disassembly, fork breakage, and joining of the broken chromosome arms. Here, we describe a mechanism for this reprogramming: the ATPase TTF2 is recruited to the replisome, where its non-catalytic N-terminal domain tethers Cyclin B-CDK1-phosphorylated TRAIP to the leading strand DNA polymerase {varepsilon} in a geometry that allows replisome ubiquitylation. Thus, a phospho-regulated architectural switch alters replisome organization in mitosis to safeguard genome integrity before chromosome segregation.

biochemistry↗

bioRxiv2024

Pipeline Olympics: continuable benchmarking of computational workflows for DNA methylation sequencing data against an experimental gold-standard

DNA methylation is a widely studied epigenetic mark and a powerful biomarker of cell type, age, environmental exposures, and disease. Whole-genome sequencing following selective conversion of unmethylated cytosines into thymines via bisulfite treatment or enzymatic methods remains the reference method for DNA methylation profiling genome-wide. While numerous software tools facilitate processing of DNA methylation sequencing reads, a comprehensive benchmarking study has been lacking thus far. In this study, we systematically compared complete computational workflows for processing DNA methylation sequencing data using a dedicated benchmarking dataset generated with five genome-wide profiling protocols. As an evaluation reference, we employed highly quantitative locus-specific measurements from our preceding benchmark of targeted DNA methylation assays. Based on this experimental gold-standard assessment and several comprehensive metrics, we identified workflows that consistently demonstrated superior performance and revealed major workflow development trends. To facilitate the sustainability of our benchmark, we implemented an interactive workflow execution and data presentation platform, adaptable to user-defined criteria and seamlessly expandable to future software.

bioinformatics↗

bioRxiv2024

In vivo analysis of the relationship between CP26 and qE-type NPQ via higher-order Arabidopsis cp26 mutants

CP26 is a monomeric minor light-harvesting complex of PSII (LHCII) protein that connects major LHCII trimers to the PSII core in photosynthetic thylakoid membranes. Previous studies have proposed that CP26 is not only involved in light harvesting but could also be involved in non-photochemical quenching (NPQ). Here, we analyzed higher-order Arabidopsis cp26 mutants using biophysical and pharmacological approaches to investigate the nature of NPQ and its relationship to known NPQ regulators (PSII subunit S (PsbS), the xanthophyll-converting enzyme VDE and the pH gradient across the thylakoid membrane). Maximum PSII quantum efficiencies (Fv/Fm) and chlorophyll fluorescence lifetimes in the dark were significantly lower in cp26 mutants, confirming that CP26 deficiency leads to a sustained quenched state even in the absence of light. Destabilized PSII-LHCII supercomplexes as observed with native PAGE analysis are the likely cause for this pre-quenched state, without other antenna proteins being able to rescue this phenotype. Further analyses revealed that cp26 mutants exhibit modest (single mutant) to highly significant (double mutants) reductions in overall NPQ capacity, which do not directly rely on PsbS and VDE (although the effect is more pronounced when these qE components are altered) but depend on thylakoid lumen acidification and protonation of protein residues. Together, these results show that the NPQ component lacking in cp26 mutants acts independently of qE and qZ and is induced in a slower phase of NPQ induction that most likely relies on pH-dependent conformational changes.

plant biology↗

bioRxiv2024

Microbial consortia driving lignocellulose transformation in agricultural woodchip bioreactors

Freshwater ecosystems can be largely affected by neighboring agriculture fields where potential fertilizer nitrate run-off may leach into surrounding water bodies. To counteract this eutrophic driver, farmers in certain areas are utilizing denitrifying woodchip bioreactors (WBRs) in which a consortium of microorganisms convert the nitrate into nitrogen-gases in anoxia, fueled by the degradation of lignocellulose. Polysaccharide-degrading strategies have been well-described for various aerobic and anaerobic systems, including the use of carbohydrate-active enzymes, utilization of lytic polysaccharide monooxygenases (LPMOs) and other redox enzymes, as well as the use of cellulosomes and polysaccharide utilization loci (PULs). However, for denitrifying microorganisms, the lignocellulose-degrading strategies remain largely unknown. Here, we have applied a combination of enrichment techniques, gas measurements, multi-omics approaches, and amplicon sequencing of fungal ITS and procaryotic 16S rRNA genes to identify microbial drivers for lignocellulose transformation in woodchip bioreactors, and their active enzymes. Our findings highlight a microbial community enriched for lignocellulose-degrading denitrifiers with key players from Giesbergeria, Cellulomonas, Azonexus, and UBA5070 (Fibrobacterota). A wide substrate specificity is observed among the many expressed carbohydrate active enzymes (CAZymes) including PULs from Bacteroidetes. This suggests a broad degradation of lignocellulose subfractions, even including enzymes with auxiliary activities whose functionality is still puzzling under strict anaerobic conditions. ImportanceFreshwater ecosystems face significant threats from agricultural runoff, which can lead to eutrophication and subsequent degradation of water quality. One solution to mitigate this issue is using denitrifying woodchip bioreactors (WBRs), where microorganisms convert nitrate into nitrogen gases utilizing lignocellulose as a carbon source. Despite the well-documented polysaccharide-degrading strategies in various systems, the mechanisms employed by denitrifying microorganisms in WBRs remain largely unexplored. This study fills a critical knowledge gap by revealing the degrading strategies of denitrifying microbial communities in WBRs. By integrating state-of-the-art techniques, we have identified key microbial drivers including Giesbergeria, Cellulomonas, Azonexus, and UBA5070 (Fibrobacterota) playing significant roles in lignocellulose transformation and showcases a broad substrate specificity and complex metabolic capability. Our findings advance the understanding of microbial ecology in WBRs and by revealing the enzymatic activities, this research may inform efforts to improving water quality, protecting aquatic ecosystems, and reducing greenhouse gas emissions from WBRs.

microbiology↗

bioRxiv2023

The genetic basis of dynamic non-photochemical quenching and photosystem II efficiency in fluctuating light reveals novel molecular targets for maize (Zea mays) improvement

Maize (Zea mays L.) is a major global crop species which uses C4 photosynthesis. Although C4 is typically considered to be more efficient than C3 photosynthesis, especially under warmer and drier conditions, there is substantial evidence that its efficiency can still be further improved, which may benefit crop performance. Improving photosynthetic efficiency via targeted manipulation of non-photochemical quenching has focused on a limited set of genes that are known to be important determinants of the NPQ response in C3 plants. The C4 pathway may alter NPQ responses but only relatively few studies have explored genetic variation in NPQ kinetics in species that perform C4 photosynthesis. In addition, studies of NPQ responses in field-grown plants of either C3 or C4 species are especially limited. Here we apply high-definition phenotyping of NPQ responses and photosynthetic efficiency and quantitative trait locus (QTL) mapping using a field-grown maize Multi-parent Advanced Generation Inter-Cross (MAGIC) population, which combines the allelic diversity of eight contrasting inbred lines. We find substantial and consistent variation for dynamic NPQ and PSII efficiency for two subsequent field seasons. Further exploration of candidate genes within three major QTL regions identified a strong impact of allelic variation in expression of the minor PSII antenna protein CP24 (LHCB6) on a major QTL for NPQ and efficiency of PSII photochemistry on chromosome 10.

plant biology↗

bioRxiv2023

Illuminating stomatal responses to red light: establishing the role of Ci-dependent vs -independent mechanisms

The stomatal response to red light appears to link stomatal conductance (gs) with photosynthetic rates. Initially, it was suggested that changes in intercellular CO 2 (Ci) provide the main cue via a Ci- dependent response. However, evidence for Ci-independent mechanisms suggests an additional, more direct relationship with photosynthesis. While both Ci-dependent and -independent mechanisms clearly function in stomatal red-light responses, little is known regarding their relative contribution. The present study aimed to quantify the relative magnitude of Ci-dependent and - independent mechanisms on the stomatal red-light response, to characterise their interplay and to assess the putative link between plastoquinone (PQ) redox state and Ci-independent stomatal responses. Red light response curves measured at a range of Ci values for wild-type Arabidopsis thaliana (Col-0) and the CO2 hyposensitive mutant, ca1ca4, allowed deconvolution of Ci-dependent and -independent pathways. Surprisingly, we observed that both mechanisms contribute equally to stomatal red-light responses, but Ci-independent stomatal opening is suppressed at high Ci. The present data are also consistent with the involvement of PQ redox in coordinating Ci-independent stomatal movements. Overall, it seems that while Ci-independent mechanisms are distinct from responses to Ci, interplay between these two pathways is important to facilitate effective coordination between gs and photosynthesis. HighlightBy controlling intercellular CO2 (Ci) at a range of values across red-light response curves, we demonstrate independent and interactive roles of Ci-dependent and -independent pathways in coordinating stomatal red-light responses.

plant biology↗

bioRxiv2023

Refined movement analysis in the Staircase test reveals differential motor deficits in mouse models of stroke

Accurate assessment of post-stroke deficits is vital in translational research. Recent advances in machine learning provide unprecedented precision in quantifying rodent motor behavior post-stroke. However, the extent to which these tools can detect lesion-specific upper extremity deficits remains unclear. Using proximal middle cerebral artery occlusion (MCAO) and cortical photothrombosis (PT), we assessed post-stroke impairments in mice through the Staircase test. Lesion locations were identified using 7T-MRI. Machine learning was applied to reconstruct kinematic trajectories using MouseReach, a data-processing toolbox. This yielded 30 refined outcome parameters effectively capturing motor deficits. Lesion reconstructions located ischemic centers in the striatum (MCAO) and sensorimotor cortex (PT). Pellet retrieval was altered in both cases but did not correlate with stroke volume or ischemia extent. Instead, cortical ischemia was characterized by increased hand slips and modified reaching success. Striatal ischemia led to progressively prolonged reach durations, mirroring delayed symptom onset in basal ganglia strokes. In summary, refined machine learning-based movement analysis revealed specific deficits in mice after cortical and striatal ischemia. These findings emphasize the importance of thorough behavioral profiling in preclinical stroke research to increase translational validity of behavioral assessments.

neuroscience↗

bioRxiv2023

EpiSegMix: A Flexible Distribution Hidden Markov Model with Duration Modeling for Chromatin State Discovery

MotivationAutomated chromatin segmentation based on ChIP-seq data reveals insights into the epigenetic regulation of chromatin accessibility. Existing segmentation methods are constrained by simplifying modeling assumptions, which may have a negative impact on the segmentation quality. ResultsWe introduce EpiSegMix, a novel segmentation method based on a hidden Markov model with flexible read count distribution types and state duration modeling, allowing for a more flexible modeling of both histone signals and segment lengths. In a comparison with two existing tools, ChromHMM, Segway and EpiCSeg, we show that EpiSegMix is more predictive of cell biology, such as gene expression. Its flexible framework enables it to fit an accurate probabilistic model, which has the potential to increase the biological interpretability of chromatin states. Availability and implementationSource code: https://gitlab.com/rahmannlab/episegmix.

bioinformatics↗

bioRxiv2023

Mutual regulation of transcriptomes between pneumocytes and fibroblasts mediates alveolar regeneration

Alveolar type 2 (AT2) and club cells are part of the stem cell niche of the lung and their differentiation is required for pulmonary homeostasis and tissue regeneration. A disturbed crosstalk between fibroblasts and epithelial cells contributes to the loss of lung structure in chronic lung diseases. Therefore, it is important to understand how fibroblasts and lung epithelial cells interact during regeneration. Here we analyzed the interaction of fibroblasts and the alveolar epithelium modelled in air-liquid interface cultures. Single-cell transcriptomics showed that co-cultivation with fibroblasts leads to increased expression of type 2 markers in pneumocytes, activation of regulons associated with maintenance of alveolar type 2 cells, and trans-differentiation of club cells towards pneumocytes. This was accompanied by an intensified transepithelial barrier. Vice versa, activation of NF{kappa}B pathways and the CEBPB regulon as well as the expression of IL-6 and other differentiation factors (e.g. FGFs) were increased in fibroblasts co-cultured with epithelial cells. Recombinant IL-6 enhanced epithelial barrier formation. Therefore, in our co-culture model, regulatory loops were identified by which lung epithelial cells mediate regeneration and differentiation of the alveolar epithelium in a cooperative manner with the mesenchymal compartment.

cell biology↗

bioRxiv2023

Molecular control of endurance training adaptation in mouse skeletal muscle

Skeletal muscle has an enormous plastic potential to adapt to various external and internal perturbations. While morphological changes in endurance-trained muscles are well-described, the molecular underpinnings of training adaptation are poorly understood. We aimed at defining the molecular signature of a trained muscle and unraveling the training statusdependent responses to an acute bout of exercise. Our results reveal that even though at baseline, the transcriptomes of trained and untrained muscles are very similar, training status substantially affects the transcriptional response to an acute challenge, both quantitatively and qualitatively, in part mediated by epigenetic modifications. Second, proteomic changes were elicited by different transcriptional modalities. Finally, transiently activated factors such as the peroxisome proliferator-activated receptor {gamma} coactivator 1 (PGC-1) are indispensable for normal training adaptation. Together, these results provide a molecular framework of the temporal and training status-dependent exercise response that defines muscle plasticity in training. HIGHLIGHTSO_LIVery few persistent transcriptional events define the trained muscle. C_LIO_LIThe training status determines the acute exercise response. C_LIO_LIEpigenetic changes shape the transcriptional specification of trained muscle. C_LIO_LIAbsence of the key regulator PGC-1 causes suboptimal training adaptations. C_LI

physiology↗

bioRxiv2023

Age-associated changes in endothelial transcriptome and chromatin landscape correlate with elevated risk of hemorrhage

Intracerebral hemorrhage (ICH), a devastating form of stroke, is a leading global cause of human death and disability. The major risk factors for ICH include increasing age, hypertension and cerebral amyloid angiopathy. Despite high mortality and morbidity associated with ICH, the mechanisms leading to blood-brain barrier (BBB) dysfunction with age and development of hemorrhagic stroke is poorly understood. In the vasculature of the central nervous system, endothelial cells (ECs) constitute the core component of the BBB and provide a physical barrier due to tight junctions, adherens junctions, and basement membrane layers. In this study, we show in brains of mice that incidents of intracerebral bleeding increase with advancing age. After isolation of an enriched population of cerebral ECs, we studied gene expression in ECs isolated from murine brains of increasing ages of 2, 6, 12, 18, and 24 months. The study reveals agedependent dysregulation of 1388 genes in the ECs, including many involved in the maintenance of BBB and vascular integrity. Since epigenetic mechanisms regulate gene expression, we also investigated age-dependent changes at the levels of CpG methylation and accessible chromatin in cerebral ECs. Our study reveals correlations between age-dependent changes in chromatin structure and gene expression. We find significant age-dependent downregulation of the apelin receptor (Aplnr) gene along with an age-dependent reduction in chromatin accessibility of the promoter of this gene. Aplnr is known to play a crucial role in positive regulation of vasodilation and is implicated in vascular health. Interestingly, we also observe an age-dependent reduction in the protein expression levels of the apelin receptor in the brain, potentially implicating the apelin receptor to be critical for the increased risk of intracerebral hemorrhage with ageing.

molecular biology↗

bioRxiv2022

A unique H2B acetylation signature marks active enhancers and predicts their target genes

Chromatin features are widely used for genome-scale mapping of enhancers. However, discriminating active enhancers from other cis-regulatory elements, predicting enhancer strength, and identifying their target genes remains challenging. Here we establish histone H2B N-terminus multisite lysine acetylation (H2BNTac) as a genuine signature of active enhancers. H2BNTac prominently marks candidate active enhancers and their target promoters and discriminates them from ubiquitously active promoters. Two mechanisms afford the distinct H2BNTac specificity. (1) Unlike H3K27ac, H2BNTac is specifically catalyzed by CBP/p300. (2) H2A-H2B, but not H3-H4, are rapidly exchanged through transcription-induced nucleosome remodeling. H2BNTac-positive candidate enhancers show a high validation rate in orthogonal enhancer activity assays, and a vast majority of endogenously active enhancers are marked by H2BNTac and H3K27ac. Notably, H2BNTac intensity predicts enhancer strength and outperforms the current state-of-the-art models in predicting enhancer target genes. These findings have broad implications for generating fine-grained enhancer maps and modeling enhancer-dependent gene regulation.

genomics↗

bioRxiv2022

The metabolic influence of the core ciliate Entodinium caudatum within the rumen microbiome

Protozoa comprise a major fraction of the microbial biomass in the rumen microbiome, of which the entodiniomorphs (order: Entodiniomorphida) and holotrichs (order: Vestibuliferida) are consistently observed to be dominant across a diverse genetic and geographical range of ruminant hosts. Despite the apparent core role that protozoal species exert, their major biological and metabolic contributions to rumen function remain largely undescribed in vivo. Here, we have leveraged (meta)genome-centric metaproteomes from rumen fluid samples originating from both cattle and goats fed diets with varying inclusion levels of lipids and starch, to detail the specific metabolic niches that protozoa occupy in the context of their microbial co-habitants. Initial proteome estimations via total protein counts and label-free quantification highlight that entodiniomorph species Entodinium and Epidinium as well as the holotrichs Dasytricha and Isotricha comprise an extensive fraction of the total rumen metaproteome. Proteomic detection of protozoal metabolism such as hydrogenases (Dasytricha, Isotricha, Epidinium, Enoploplastron), carbohydrate-active enzymes (Epidinium, Diplodinium, Enoploplastron, Polyplastron), microbial predation (Entodinium) and volatile fatty acid production (Entodinium and Epidinium) was observed at increased levels in high methane-emitting animals. Despite certain protozoal species having well-established reputations for digesting starch, they were unexpectedly less detectable in low methane emitting-animals fed high starch diets, which were instead dominated by propionate/succinate-producing bacterial populations suspected of being resistant to predation irrespective of host. Finally, we reaffirmed our abovementioned observations in geographically independent datasets, thus illuminating the substantial metabolic influence that under-explored eukaryotic populations have in the rumen, with greater implications for both digestion and methane metabolism.

microbiology↗