Phylogeny, morphology, virulence, ecology, and host range of Ordospora pajunii (Ordosporidae), a microsporidian symbiont of Daphnia spp.

The impacts of microsporidia on host individuals are frequently subtle and can be context dependent. A key example of the latter comes from a recently discovered microsporidian symbiont of Daphnia, the net impact of which was found to shift from negative to positive based on environmental context. Given this, we hypothesized low baseline virulence of the microsporidian; here, we investigated the impact of infection on hosts in controlled conditions and the absence of other stressors. We also investigated its phylogenetic position, ecology, and host range. The genetic data indicate that the symbiont is Ordospora pajunii, a newly described microsporidian parasite of Daphnia. We show that O. pajunii infection damages the gut, causing infected epithelial cells to lose microvilli and then rupture. The prevalence of this microsporidian could be high (up to 100% in the lab and 77% of adults in the field). Its overall virulence was low in most cases, but some genotypes suffered reduced survival and/or reproduction. Susceptibility and virulence were strongly host-genotype dependent. We found that North American O. pajunii were able to infect multiple Daphnia species, including the European species Daphnia longispina, as well as Ceriodaphnia spp. Given the low, often undetectable virulence of this microsporidian and potentially far-reaching consequences of infections for the host when interacting with other pathogens or food, this Daphnia-O. pajunii symbiosis emerges as a valuable system for studying the mechanisms of context-dependent shifts between mutualism and parasitism, as well as for understanding how symbionts might alter host interactions with resources. IMPORTANCE: The net outcome of symbiosis depends on the costs and benefits to each partner. Those can be context dependent, driving the potential for an interaction to change between parasitism and mutualism. Understanding the baseline fitness impact in an interaction can help us understand those shifts; for an organism that is generally parasitic, it should be easier for it to become a mutualist if its baseline virulence is relatively low. Recently, a microsporidian was found to become beneficial to its Daphnia hosts in certain ecological contexts, but little was known about the symbiont (including its species identity). Here, we identify it as the microsporidium Ordospora pajunii. Despite the parasitic nature of microsporidia, we found O. pajunii to be, at most, mildly virulent; this helps explain why it can shift toward mutualism in certain ecological contexts and helps establish O. pajunii is a valuable model for investigating shifts along the mutualism-parasitism continuum.

Potential for functional divergence in ectomycorrhizal fungal communities across a precipitation gradient.

Functional traits influence the assembly of microbial communities, but identifying these traits in the environment has remained challenging. We studied ectomycorrhizal fungal (EMF) communities inhabiting Populus trichocarpa roots distributed across a precipitation gradient in the Pacific Northwest, USA. We profiled these communities using taxonomic (meta-barcoding) and functional (metagenomic) approaches. We hypothesized that genes involved in fungal drought-stress tolerance and fungal mediated plant water uptake would be most abundant in drier soils. We were unable to detect support for this hypothesis; instead, the abundance of genes involved in melanin synthesis, hydrophobins, aquaporins, trehalose-synthases, and other gene families exhibited no significant shifts across the gradient. Finally, we studied variation in sequence homology for certain genes, finding that fungal communities in dry soils are composed of distinct aquaporin and hydrophobin gene sequences. Altogether, our results suggest that while EMF communities exhibit significant compositional shifts across this gradient, coupled functional turnover, at least as inferred using community metagenomics is limited. Accordingly, the consequences of these distinct EMF communities on plant water uptake remain critically unknown, and future studies targeting the expression of genes involved in drought stress tolerance are required.

Draft genome sequence of Yarrowia lipolytica NRRL Y-64008, an oleaginous yeast capable of growing on lignocellulosic hydrolysates.

Yarrowia lipolytica is an oleaginous yeast that produces high titers of fatty acid-derived biofuels and biochemicals. It can grow on hydrophobic carbon sources and lignocellulosic hydrolysates. The genome sequence of Y. lipolytica NRRL Y-64008 is reported to aid in its development as a biotechnological chassis for producing biofuels and bioproducts.

Near-complete genome sequence of Lipomyces tetrasporous NRRL Y-64009, an oleaginous yeast capable of growing on lignocellulosic hydrolysates.

Lipomyces tetrasporous is an oleaginous yeast that can utilize a variety of plant-based sugars. It accumulates lipids during growth on lignocellulosic biomass hydrolysates. We present the annotated genome sequence of L. tetrasporous NRRL Y-64009 to aid in its development as a platform organism for producing lipids and lipid-based bioproducts.

A scaffolded and annotated reference genome of giant kelp (Macrocystis pyrifera).

Macrocystis pyrifera (giant kelp), is a brown macroalga of great ecological importance as a primary producer and structure-forming foundational species that provides habitat for hundreds of species. It has many commercial uses (e.g. source of alginate, fertilizer, cosmetics, feedstock). One of the limitations to exploiting giant kelp's economic potential and assisting in giant kelp conservation efforts is a lack of genomic tools like a high quality, contiguous reference genome with accurate gene annotations. Reference genomes attempt to capture the complete genomic sequence of an individual or species, and importantly provide a universal structure for comparison across a multitude of genetic experiments, both within and between species. We assembled the giant kelp genome of a haploid female gametophyte de novo using PacBio reads, then ordered contigs into chromosome level scaffolds using Hi-C. We found the giant kelp genome to be 537 MB, with a total of 35 scaffolds and 188 contigs. The assembly N50 is 13,669,674 with GC content of 50.37%. We assessed the genome completeness using BUSCO, and found giant kelp contained 94% of the BUSCO genes from the stramenopile clade. Annotation of the giant kelp genome revealed 25,919 genes. Additionally, we present genetic variation data based on 48 diploid giant kelp sporophytes from three different Southern California populations that confirms the population structure found in other studies of these populations. This work resulted in a high-quality giant kelp genome that greatly increases the genetic knowledge of this ecologically and economically vital species.

Terabase-Scale Coassembly of a Tropical Soil Microbiome.

Petabases of environmental metagenomic data are publicly available, presenting an opportunity to characterize complex environments and discover novel lineages of life. Metagenome coassembly, in which many metagenomic samples from an environment are simultaneously analyzed to infer the underlying genomes' sequences, is an essential tool for achieving this goal. We applied MetaHipMer2, a distributed metagenome assembler that runs on supercomputing clusters, to coassemble 3.4 terabases (Tbp) of metagenome data from a tropical soil in the Luquillo Experimental Forest (LEF), Puerto Rico. The resulting coassembly yielded 39 high-quality (>90% complete, <5% contaminated, with predicted 23S, 16S, and 5S rRNA genes and ≥18 tRNAs) metagenome-assembled genomes (MAGs), including two from the candidate phylum Eremiobacterota. Another 268 medium-quality (≥50% complete, <10% contaminated) MAGs were extracted, including the candidate phyla Dependentiae, Dormibacterota, and Methylomirabilota. In total, 307 medium- or higher-quality MAGs were assigned to 23 phyla, compared to 294 MAGs assigned to nine phyla in the same samples individually assembled. The low-quality (<50% complete, <10% contaminated) MAGs from the coassembly revealed a 49% complete rare biosphere microbe from the candidate phylum FCPU426 among other low-abundance microbes, an 81% complete fungal genome from the phylum Ascomycota, and 30 partial eukaryotic MAGs with ≥10% completeness, possibly representing protist lineages. A total of 22,254 viruses, many of them low abundance, were identified. Estimation of metagenome coverage and diversity indicates that we may have characterized ≥87.5% of the sequence diversity in this humid tropical soil and indicates the value of future terabase-scale sequencing and coassembly of complex environments. IMPORTANCE Petabases of reads are being produced by environmental metagenome sequencing. An essential step in analyzing these data is metagenome assembly, the computational reconstruction of genome sequences from microbial communities. "Coassembly" of metagenomic sequence data, in which multiple samples are assembled together, enables more complete detection of microbial genomes in an environment than "multiassembly," in which samples are assembled individually. To demonstrate the potential for coassembling terabases of metagenome data to drive biological discovery, we applied MetaHipMer2, a distributed metagenome assembler that runs on supercomputing clusters, to coassemble 3.4 Tbp of reads from a humid tropical soil environment. The resulting coassembly, its functional annotation, and analysis are presented here. The coassembly yielded more, and phylogenetically more diverse, microbial, eukaryotic, and viral genomes than the multiassembly of the same data. Our resource may facilitate the discovery of novel microbial biology in tropical soils and demonstrates the value of terabase-scale metagenome sequencing.

An HMM approach expands the landscape of sesquiterpene cyclases across the kingdom Fungi.

Sesquiterpene cyclases (STC) catalyse the cyclization of the C15 molecule farnesyl diphosphate into a vast variety of mono- or polycyclic hydrocarbons and, for a few enzymes, oxygenated structures, with diverse stereogenic centres. The huge diversity in sesquiterpene skeleton structures in nature is primarily the result of the type of cyclization driven by the STC. Despite the phenomenal impact of fungal sesquiterpenes on the ecology of fungi and their potentials for applications, the fungal sesquiterpenome is largely untapped. The identification of fungal STC is generally based on protein sequence similarity with characterized enzymes. This approach has improved our knowledge on STC in a few fungal species, but it has limited success for the discovery of distant sequences. Besides, the tools based on secondary metabolite biosynthesis gene clusters have shown poor performance for terpene cyclases. Here, we used four sets of sequences of fungal STC that catalyse four types of cyclization, and specific amino acid motives to identify phylogenetically related sequences in the genomes of basidiomycetes fungi from the order Polyporales. We validated that four STC genes newly identified from the genome sequence of Leiotrametes menziesii, each classified in a different phylogenetic clade, catalysed a predicted cyclization of farnesyl diphosphate. We built HMM models and searched STC genes in 656 fungal genomes genomes. We identified 5605 STC genes, which were classified in one of the four clades and had a predicted cyclization mechanism. We noticed that the HMM models were more accurate for the prediction of the type of cyclization catalysed by basidiomycete STC than for ascomycete STC.

Dataset of 143 metagenome-assembled genomes from the Arctic and Atlantic Oceans, including 21 for eukaryotic organisms.

This article presents metagenome-assembled genomes (MAGs) for both eukaryotic and prokaryotic organisms originating from the Arctic and Atlantic oceans, along with gene prediction and functional annotation for MAGs from both domains. Eleven samples from the chlorophyll-a maximum layer of the surface ocean were collected during two cruises in 2012; six from the Arctic in June-July on ARK-XXVII/1 (PS80), and five from the Atlantic in November on ANT-XXIX/1 (PS81). Sequencing and assembly was carried out by the Joint Genome Institute (JGI), who provide annotation of the assembled sequences, and 122 MAGs for prokaryotic organisms. A subsequent binning process identified 21 MAGs for eukaryotic organisms, mostly identified as Mamiellophyceae or Bacillariophyceae. The data for each MAG includes sequences in FASTA format, and tables of functional annotation of genes. For eukaryotic MAGs, transcript and protein sequences for predicted genes are available. A spreadsheet is provided summarising quality measures and taxonomic classifications for each MAG. These data provide draft genomes for uncultured marine microbes, including some of the first MAGs for polar eukaryotes, and can provide reference genetic data for these environments, or used in genomics-based comparison between environments.

Two major chromosome evolution events with unrivaled conserved gene content in pomegranate.

Pomegranate has a unique evolutionary history given that different cultivars have eight or nine bivalent chromosomes with possible crossability between the two classes. Therefore, it is important to study chromosome evolution in pomegranate to understand the dynamics of its population. Here, we de novo assembled the Azerbaijani cultivar "Azerbaijan guloyshasi" (AG2017; 2n = 16) and re-sequenced six cultivars to track the evolution of pomegranate and to compare it with previously published de novo assembled and re-sequenced cultivars. High synteny was observed between AG2017, Bhagawa (2n = 16), Tunisia (2n = 16), and Dabenzi (2n = 18), but these four cultivars diverged from the cultivar Taishanhong (2n = 18) with several rearrangements indicating the presence of two major chromosome evolution events. Major presence/absence variations were not observed as >99% of the five genomes aligned across the cultivars, while >99% of the pan-genic content was represented by Tunisia and Taishanhong only. We also revisited the divergence between soft- and hard-seeded cultivars with less structured population genomic data, compared to previous studies, to refine the selected genomic regions and detect global migration routes for pomegranate. We reported a unique admixture between soft- and hard-seeded cultivars that can be exploited to improve the diversity, quality, and adaptability of local pomegranate varieties around the world. Our study adds body knowledge to understanding the evolution of the pomegranate genome and its implications for the population structure of global pomegranate diversity, as well as planning breeding programs aiming to develop improved cultivars.

Divergent Evolution of Early Terrestrial Fungi Reveals the Evolution of Mucormycosis Pathogenicity Factors.

Fungi have evolved over millions of years and their species diversity is predicted to be the second largest on the earth. Fungi have cross-kingdom interactions with many organisms that have mutually shaped their evolutionary trajectories. Zygomycete fungi hold a pivotal position in the fungal tree of life and provide important perspectives on the early evolution of fungi from aquatic to terrestrial environments. Phylogenomic analyses have found that zygomycete fungi diversified into two separate clades, the Mucoromycota which are frequently associated with plants and Zoopagomycota that are commonly animal-associated fungi. Genetic elements that contributed to the fitness and divergence of these lineages may have been shaped by the varied interactions these fungi have had with plants, animals, bacteria, and other microbes. To investigate this, we performed comparative genomic analyses of the two clades of zygomycetes in the context of Kingdom Fungi, benefiting from our generation of a new collection of zygomycete genomes, including nine produced for this study. We identified lineage-specific genomic content that may contribute to the disparate biology observed in these zygomycetes. Our findings include the discovery of undescribed diversity in CotH, a Mucormycosis pathogenicity factor, which was found in a broad set of zygomycetes. Reconciliation analysis identified multiple duplication events and an expansion of CotH copies throughout the Mucoromycotina, Mortierellomycotina, Neocallimastigomycota, and Basidiobolus lineages. A kingdom-level phylogenomic analysis also identified new evolutionary relationships within the subphyla of Mucoromycota and Zoopagomycota, including supporting the sister-clade relationship between Glomeromycotina and Mortierellomycotina and the placement of Basidiobolus as sister to other Zoopagomycota lineages.

Genetic and Structural Diversity of Prokaryotic Ice-Binding Proteins from the Central Arctic Ocean.

Ice-binding proteins (IBPs) are a group of ecologically and biotechnologically relevant enzymes produced by psychrophilic organisms. Although putative IBPs containing the domain of unknown function (DUF) 3494 have been identified in many taxa of polar microbes, our knowledge of their genetic and structural diversity in natural microbial communities is limited. Here, we used samples from sea ice and sea water collected in the central Arctic Ocean as part of the MOSAiC expedition for metagenome sequencing and the subsequent analyses of metagenome-assembled genomes (MAGs). By linking structurally diverse IBPs to particular environments and potential functions, we reveal that IBP sequences are enriched in interior ice, have diverse genomic contexts and cluster taxonomically. Their diverse protein structures may be a consequence of domain shuffling, leading to variable combinations of protein domains in IBPs and probably reflecting the functional versatility required to thrive in the extreme and variable environment of the central Arctic Ocean.

Convergent evolution and horizontal gene transfer in Arctic Ocean microalgae.

Microbial communities in the world ocean are affected strongly by oceanic circulation, creating characteristic marine biomes. The high connectivity of most of the ocean makes it difficult to disentangle selective retention of colonizing genotypes (with traits suited to biome specific conditions) from evolutionary selection, which would act on founder genotypes over time. The Arctic Ocean is exceptional with limited exchange with other oceans and ice covered since the last ice age. To test whether Arctic microalgal lineages evolved apart from algae in the global ocean, we sequenced four lineages of microalgae isolated from Arctic waters and sea ice. Here we show convergent evolution and highlight geographically limited HGT as an ecological adaptive force in the form of PFAM complements and horizontal acquisition of key adaptive genes. Notably, ice-binding proteins were acquired and horizontally transferred among Arctic strains. A comparison with Tara Oceans metagenomes and metatranscriptomes confirmed mostly Arctic distributions of these IBPs. The phylogeny of Arctic-specific genes indicated that these events were independent of bacterial-sourced HGTs in Antarctic Southern Ocean microalgae.

A Remarkable Expansion of Oligopeptide Transporter Genes in Rust Fungi (Pucciniales) Suggests a Specialization in Nutrient Acquisition for Obligate Biotrophy.

Nutrient acquisition by rust fungi during their biotrophic growth has been assigned to a few transporters expressed in haustorial infection structures. We performed a comparative genomic analysis of all transporter genes (hereafter termed transportome) classified according to the Transporter Classification Database, focusing specifically on rust fungi (order Pucciniales) versus other species in the Dikarya. We also surveyed expression of transporter genes in the poplar rust fungus for which transcriptomics data are available across the whole life cycle. Despite a significant increase in gene number, rust fungi presented a reduced transportome compared with most fungi in the Dikarya. However, a few transporter families in the subclass Porters showed significant expansions. Notably, three metal transport-related families involved in the import, export, and sequestration of metals were expanded in Pucciniales and expressed at various stages of the rust life cycle, suggesting a tight regulation of metal homeostasis. The most remarkable gene expansion in the Pucciniales was observed for the oligopeptide transporter (OPT) family, with 25 genes on average compared with seven to 14 genes in the other surveyed taxonomical ranks. A phylogenetic analysis showed several specific expansion events at the root of the order Pucciniales with subsequent expansions in rust taxonomical families. The OPT genes showed dynamic expression patterns along the rust life cycle and more particularly during infection of the poplar host tree, suggesting a possible specialization for the acquisition of nitrogen and sulfur through the transport of oligopeptides from the host during biotrophic growth.

Multiomics in the central Arctic Ocean for benchmarking biodiversity change.

Multiomics approaches need to be applied in the central Arctic Ocean to benchmark biodiversity change and to identify novel species and their genes. As part of MOSAiC, EcoOmics will therefore be essential for conservation and sustainable bioprospecting in one of the least explored ecosystems on Earth.

Wildfire-dependent changes in soil microbiome diversity and function.

Forest soil microbiomes have crucial roles in carbon storage, biogeochemical cycling and rhizosphere processes. Wildfire season length, and the frequency and size of severe fires have increased owing to climate change. Fires affect ecosystem recovery and modify soil microbiomes and microbially mediated biogeochemical processes. To study wildfire-dependent changes in soil microbiomes, we characterized functional shifts in the soil microbiota (bacteria, fungi and viruses) across burn severity gradients (low, moderate and high severity) 1 yr post fire in coniferous forests in Colorado and Wyoming, USA. We found severity-dependent increases of Actinobacteria encoding genes for heat resistance, fast growth, and pyrogenic carbon utilization that might enhance post-fire survival. We report that increased burn severity led to the loss of ectomycorrhizal fungi and less tolerant microbial taxa. Viruses remained active in post-fire soils and probably influenced carbon cycling and biogeochemistry via turnover of biomass and ecosystem-relevant auxiliary metabolic genes. Our genome-resolved analyses link post-fire soil microbial taxonomy to functions and reveal the complexity of post-fire soil microbiome activity.

Near-Complete Genome Sequence of Zygosaccharomyces rouxii NRRL Y-64007, a Yeast Capable of Growing on Lignocellulosic Hydrolysates.

The halotolerant and osmotolerant yeast Zygosaccharomyces rouxii can produce multiple volatile compounds and has the ability to grow on lignocellulosic hydrolysates. We report the annotated genome sequence of Z. rouxii NRRL Y-64007 to support its development as a platform organism for biofuel and bioproduct production.

Metagenome-assembled genomes of phytoplankton microbiomes from the Arctic and Atlantic Oceans.

BACKGROUND: Phytoplankton communities significantly contribute to global biogeochemical cycles of elements and underpin marine food webs. Although their uncultured genomic diversity has been estimated by planetary-scale metagenome sequencing and subsequent reconstruction of metagenome-assembled genomes (MAGs), this approach has yet to be applied for complex phytoplankton microbiomes from polar and non-polar oceans consisting of microbial eukaryotes and their associated prokaryotes. RESULTS: Here, we have assembled MAGs from chlorophyll a maximum layers in the surface of the Arctic and Atlantic Oceans enriched for species associations (microbiomes) with a focus on pico- and nanophytoplankton and their associated heterotrophic prokaryotes. From 679 Gbp and estimated 50 million genes in total, we recovered 143 MAGs of medium to high quality. Although there was a strict demarcation between Arctic and Atlantic MAGs, adjacent sampling stations in each ocean had 51-88% MAGs in common with most species associations between Prasinophytes and Proteobacteria. Phylogenetic placement revealed eukaryotic MAGs to be more diverse in the Arctic whereas prokaryotic MAGs were more diverse in the Atlantic Ocean. Approximately 70% of protein families were shared between Arctic and Atlantic MAGs for both prokaryotes and eukaryotes. However, eukaryotic MAGs had more protein families unique to the Arctic whereas prokaryotic MAGs had more families unique to the Atlantic. CONCLUSION: Our study provides a genomic context to complex phytoplankton microbiomes to reveal that their community structure was likely driven by significant differences in environmental conditions between the polar Arctic and warm surface waters of the tropical and subtropical Atlantic Ocean. Video Abstract.

Annotation survey and life cycle transcriptomics of transcription factors in rust fungi (Pucciniales) identify a possible role for cold shock proteins in dormancy exit.

Fungi of the order Pucciniales are obligate plant biotrophs causing rust diseases. They exhibit a complex life cycle with the production of up to five spore types, infection of two unrelated hosts and an overwintering stage. Transcription factors (TFs) are key regulators of gene expression in eukaryote cells. In order to better understand genetic programs expressed during major transitions of the rust life cycle, we surveyed the complement of TFs in fungal genomes with an emphasis on Pucciniales. We found that despite their large gene numbers, rust genomes have a reduced repertoire of TFs compared to other fungi. The proportions of C2H2 and Zinc cluster - two of the most represented TF families in fungi - indicate differences in their evolutionary relationships in Pucciniales and other fungal taxa. The regulatory gene family encoding cold shock protein (CSP) showed a striking expansion in Pucciniomycotina with specific duplications in the order Pucciniales. The survey of expression profiles collected by transcriptomics along the life cycle of the poplar rust fungus revealed TF genes related to major biological transitions, e.g. response to environmental cues and host infection. Particularly, poplar rust CSPs were strongly expressed in basidia produced after the overwintering stage suggesting a possible role in dormancy exit. Expression during transition from dormant telia to basidia confirmed the specific expression of the three poplar rust CSP genes. Their heterologous expression in yeast improved cell growth after cold stress exposure, suggesting a probable regulatory function when the poplar rust fungus exits dormancy. This study addresses for the first time TF and regulatory genes involved in developmental transition in the rust life cycle opening perspectives to further explore molecular regulation in the biology of the Pucciniales.

Kingdom-Wide Analysis of Fungal Protein-Coding and tRNA Genes Reveals Conserved Patterns of Adaptive Evolution.

Protein-coding genes evolved codon usage bias due to the combined but uneven effects of adaptive and nonadaptive influences. Studies in model fungi agree on codon usage bias as an adaptation for fine-tuning gene expression levels; however, such knowledge is lacking for most other fungi. Our comparative genomics analysis of over 450 species supports codon usage and transfer RNAs (tRNAs) as coadapted for translation speed and this is most likely a realization of convergent evolution. Rather than drift, phylogenetic reconstruction inferred adaptive radiation as the best explanation for the variation of interspecific codon usage bias. Although the phylogenetic signals for individual codon and tRNAs frequencies are lower than expected by genetic drift, we found remarkable conservation of highly expressed genes being codon optimized for translation by the most abundant tRNAs, especially by inosine-modified tRNAs. As an application, we present a sequence-to-expression neural network that uses codons to reliably predict highly expressed transcripts. The kingdom Fungi, with over a million species, includes many key players in various ecosystems and good targets for biotechnology. Collectively, our results have implications for better understanding the evolutionary success of fungi, as well as informing the biosynthetic manipulation of fungal genes.

Co­cultivation of the anaerobic fungus Caecomyces churrovis with Methanobacterium bryantii enhances transcription of carbohydrate binding modules, dockerins, and pyruvate formate lyases on specific substrates.

Anaerobic fungi and methanogenic archaea are two classes of microorganisms found in the rumen microbiome that metabolically interact during lignocellulose breakdown. Here, stable synthetic co-cultures of the anaerobic fungus Caecomyces churrovis and the methanogen Methanobacterium bryantii (not native to the rumen) were formed, demonstrating that microbes from different environments can be paired based on metabolic ties. Transcriptional and metabolic changes induced by methanogen co-culture were evaluated in C. churrovis across a variety of substrates to identify mechanisms that impact biomass breakdown and sugar uptake. A high-quality genome of C. churrovis was obtained and annotated, which is the first sequenced genome of a non-rhizoid-forming anaerobic fungus. C. churrovis possess an abundance of CAZymes and carbohydrate binding modules and, in agreement with previous studies of early-diverging fungal lineages, N6-methyldeoxyadenine (6mA) was associated with transcriptionally active genes. Co-culture with the methanogen increased overall transcription of CAZymes, carbohydrate binding modules, and dockerin domains in co-cultures grown on both lignocellulose and cellulose and caused upregulation of genes coding associated enzymatic machinery including carbohydrate binding modules in family 18 and dockerin domains across multiple growth substrates relative to C. churrovis monoculture. Two other fungal strains grown on a reed canary grass substrate in co-culture with the same methanogen also exhibited high log2-fold change values for upregulation of genes encoding carbohydrate binding modules in families 1 and 18. Transcriptional upregulation indicated that co-culture of the C. churrovis strain with a methanogen may enhance pyruvate formate lyase (PFL) function for growth on xylan and fructose and production of bottleneck enzymes in sugar utilization pathways, further supporting the hypothesis that co-culture with a methanogen may enhance certain fungal metabolic functions. Upregulation of CBM18 may play a role in fungal-methanogen physical associations and fungal cell wall development and remodeling.