Engineering topics

Froula, Jeff

Publications and source records attributed to Froula, Jeff.

9 records

DOE OSTI2017

Gene and transcript abundances of bacterial type III secretion systems from the rumen microbiome are correlated with methane yield in sheep

Background: Ruminants are important contributors to global methane emissions via microbial fermentation in their reticulo-rumens. This study is part of a larger program, characterising the rumen microbiomes of sheep which vary naturally in methane yield (g CH4/kg DM/day) and aims to defne diferences in microbial communities, and in gene and transcript abundances that can explain the animal methane phenotype. Methods: Rumen microbiome metagenomic and metatranscriptomic data were analysed by Gene Set Enrichment, sparse partial least squares regression and the Wilcoxon Rank Sum test to estimate correlations between specifc KEGG bacterial pathways/genes and high methane yield in sheep. KEGG genes enriched in high methane yield sheep were reassembled from raw reads and existing contigs and analysed by MEGAN to predict their phylogenetic origin. Protein coding sequences from Succinivibrio dextrinosolvens strains were analysed using Efective DB to predict bacterial type III secreted proteins. The efect of S. dextrinosolvens strain H5 growth on methane formation by rumen methanogens was explored using co-cultures. Results: Detailed analysis of the rumen microbiomes of high methane yield sheep shows that gene and transcript abundances of bacterial type III secretion system genes are positively correlated with methane yield in sheep. Most of the bacterial type III secretion system genes could not be assigned to a particular bacterial group, but several genes were afliated with the genus Succinivibrio, and searches of bacterial genome sequences found that strains of S. dextrinosolvens were part of a small group of rumen bacteria that encode this type of secretion system. In co-culture experiments, S. dextrinosolvens strain H5 showed a growth-enhancing efect on a methanogen belonging to the order Methanomassiliicoccales, and inhibition of a representative of the Methanobrevibacter gottschalkii clade. Conclusions: This is the frst report of bacterial type III secretion system genes being associated with high methane emissions in ruminants, and identifes these secretions systems as potential new targets for methane mitigation research. The effects of S. dextrinosolvens on the growth of rumen methanogens in co-cultures indicate that bacteriamethanogen interactions are important modulators of methane production in ruminant animals.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2016

Rumen metagenome and metatranscriptome analyses of low methane yield sheep reveals a Sharpea -enriched microbiome characterised by lactic acid formation and utilisation

Enteric fermentation by farmed ruminant animals is a major source of methane and constitutes the second largest anthropogenic contributor to global warming. Reducing methane emissions from ruminants is needed to ensure sustainable animal production in the future. Methane yield varies naturally in sheep and is a heritable trait that can be used to select animals that yield less methane per unit of feed eaten. We previously demonstrated elevated expression of hydrogenotrophic methanogenesis pathway genes of methanogenic archaea in the rumens of high methane yield (HMY) sheep compared to their low methane yield (LMY) counterparts. Methane production in the rumen is strongly connected to microbial hydrogen production through fermentation processes. In this study, we investigate the contribution that rumen bacteria make to methane yield phenotypes in sheep. Using deep sequence metagenome and metatranscriptome datasets in combination with 16S rRNA gene amplicon sequencing from HMY and LMY sheep, we show enrichment of lactate-producing Sharpea spp. in LMY sheep bacterial communities. Increased gene and transcript abundances for sugar import and utilisation and production of lactate, propionate and butyrate were also observed in LMY animals. Sharpea azabuensis and Megasphaera spp. act as important drivers of lactate production and utilisation according to phylogenetic analysis and read mappings. Our findings show that the rumen microbiome in LMY animals supports a rapid heterofermentative growth, leading to lactate production. We postulate that lactate is subsequently metabolised mainly to butyrate in LMY animals, producing 2 mol of hydrogen and 0.5 mol of methane per mol hexose, which represents 24 % less than the 0.66 mol of methane formed from the 2.66 mol of hydrogen produced if hexose fermentation was directly to acetate and butyrate. These findings are consistent with the theory that a smaller rumen size with a higher turnover rate, where rapid heterofermentative growth would be an advantage, results in lower hydrogen production and lower methane formation. Together with previous methanogen gene expression data, this builds a strong concept of how animal traits and microbial communities shape the methane phenotype in sheep.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2016

The Epigenomic Landscape of Prokaryotes

DNA methylation acts in concert with restriction enzymes to protect the integrity of prokaryotic genomes. Studies in a limited number of organisms suggest that methylation also contributes to prokaryotic genome regulation, but the prevalence and properties of such non-restriction-associated methylation systems remain poorly understood. Here, we used single molecule, real-time sequencing to map DNA modifications including m6A, m4C, and m5C across the genomes of 230 diverse bacterial and archaeal species. We observed DNA methylation in nearly all (93%) organisms examined, and identified a total of 834 distinct reproducibly methylated motifs. This data enabled annotation of the DNA binding specificities of 620 DNA Methyltransferases (MTases), doubling known specificities for previously hard to study Type I, IIG and III MTases, and revealing their extraordinary diversity. Strikingly, 48% of organisms harbor active Type II MTases with no apparent cognate restriction enzyme. These active ‘orphan’ MTases are present in diverse bacterial and archaeal phyla and show motif specificities and methylation patterns consistent with functions in gene regulation and DNA replication. Our results reveal the pervasive presence of DNA methylation throughout the prokaryotic kingdoms, as well as the diversity of sequence specificities and potential functions of DNA methylation systems.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2016

Genome-wide selective sweeps and gene-specific sweeps in natural bacterial populations

Multiple models describe the formation and evolution of distinct microbial phylogenetic groups. These evolutionary models make different predictions regarding how adaptive alleles spread through populations and how genetic diversity is maintained. Processes predicted by competing evolutionary models, for example, genome-wide selective sweeps vs gene-specific sweeps, could be captured in natural populations using time-series metagenomics if the approach were applied over a sufficiently long time frame. Direct observations of either process would help resolve how distinct microbial groups evolve. Using a 9-year metagenomic study of a freshwater lake (2005–2013), we explore changes in single-nucleotide polymorphism (SNP) frequencies and patterns of gene gain and loss in 30 bacterial populations. SNP analyses revealed substantial genetic heterogeneity within these populations, although the degree of heterogeneity varied by >1000-fold among populations. SNP allele frequencies also changed dramatically over time within some populations. Interestingly, nearly all SNP variants were slowly purged over several years from one population of green sulfur bacteria, while at the same time multiple genes either swept through or were lost from this population. Furthermore, these patterns were consistent with a genome-wide selective sweep in progress, a process predicted by the ‘ecotype model’ of speciation but not previously observed in nature. In contrast, other populations contained large, SNP-free genomic regions that appear to have swept independently through the populations prior to the study without purging diversity elsewhere in the genome. Finally, evidence for both genome-wide and gene-specific sweeps suggests that different models of bacterial speciation may apply to different populations coexisting in the same environment.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2015

MetaBAT, an efficient tool for accurately reconstructing single genomes from complex microbial communities

Grouping large genomic fragments assembled from shotgun metagenomic sequences to deconvolute complex microbial communities, or metagenome binning, enables the study of individual organisms and their interactions. Because of the complex nature of these communities, existing metagenome binning methods often miss a large number of microbial species. In addition, most of the tools are not scalable to large datasets. Here we introduce automated software called MetaBAT that integrates empirical probabilistic distances of genome abundance and tetranucleotide frequency for accurate metagenome binning. MetaBAT outperforms alternative methods in accuracy and computational efficiency on both synthetic and real metagenome datasets. Lastly, it automatically forms hundreds of high quality genome bins on a very large assembly consisting millions of contigs in a matter of hours on a single node. MetaBAT is open source software and available at https://bitbucket.org/berkeleylab/metabat.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2014

Methane yield phenotypes linked to differential gene expression in the sheep rumen microbiome

Ruminant livestock represent the single largest anthropogenic source of the potent greenhouse gas methane, which is generated by methanogenic archaea residing in ruminant digestive tracts. While differences between individual animals of the same breed in the amount of methane produced have been observed, the basis for this variation remains to be elucidated. To explore the mechanistic basis of this methane production, we measured methane yields from 22 sheep, which revealed that methane yields are a reproducible, quantitative trait. Deep metagenomic and metatranscriptomic sequencing demonstrated a similar abundance of methanogens and methanogenesis pathway genes in high and low methane emitters. However, transcription of methanogenesis pathway genes was substantially increased in sheep with high methane yields. These results identify a discrete set of rumen methanogens whose methanogenesis pathway transcription profiles correlate with methane yields and provide new targets for CH4 mitigation at the levels of microbiota composition and transcriptional regulation.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2014

MetaBAT

Assembling individual genomes from shotgun metagenomic sequences derived from complex microbial communities is so far one of the most challenging problems in bioinformatics.As it is impractical to directly assemble full-length genomes, a first step that groups contigs from the same organisms, called metagenome binning, has been developed to provide insights of individual organisms. However, current binning methods perform poorly in the context of large complex community, and as a result they fail to recover many novel genomes. To overcome this limitation, we developed integrated software, called MetaBAT, which automatically forms hundreds of individual genome bins from metagenome contigs. Probabilistic models of abundance and tetranucleotide frequency were trained by extensive empirical studies and integrated to decide the membership of contigs iteratively. To test the performance of MetaBAT, we applied MetaBAT to both synthetic and several large-scale real world metagenome datasets. By using two independent metrics, we demonstrate that in all the data sets tested MetaBAT achieves good sensitivity (16~87%) and very high specificity (56~99%) in forming genome bins. Further analyses of the novel genomes recovered from the human gut microbiome suggest a subset of these genomes are potentially associated with pathological conditions. In conclusion, we believe MetaBAT is a powerful tool

Kang, Dongwan↗

DOE OSTI2014

MetaBAT: Metagenome Binning based on Abundance and Tetranucleotide frequence

Grouping large fragments assembled from shotgun metagenomic sequences to deconvolute complex microbial communities, or metagenome binning, enables the study of individual organisms and their interactions. Here we developed automated metagenome binning software, called MetaBAT, which integrates empirical probabilistic distances of genome abundance and tetranucleotide frequency. On synthetic datasets MetaBAT on average achieves 98percent precision and 90percent recall at the strain level with 281 near complete unique genomes. Applying MetaBAT to a human gut microbiome data set we recovered 176 genome bins with 92percent precision and 80percent recall. Further analyses suggest MetaBAT is able to recover genome fragments missed in reference genomes up to 19percent, while 53 genome bins are novel. In summary, we believe MetaBAT is a powerful tool to facilitate comprehensive understanding of complex microbial communities.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2014

Dynamics of Sequence -Discrete Bacterial Populations Inferred Using Metagenomes

From a multi-year metagenomic time series of two dissimilar Wisconsin lakes we have assembled dozens of genomes using a novel approach that bins contigs into distinct genome based on sequence composition, e.g. kmer frequencies, and contig coverage patterns at various times points. Next, we investigated how these genomes, which represent sequence-discrete bacterial populations, evolved over time and used the time series to discover the population dynamics. For example, we explored changes in single nucleotide polymorphism (SNP) frequencies as well as patterns of gene gain and loss in multiple populations. Interestingly, SNP diversity was purged at nearly every genome position in some populations during the course of this study, suggesting these populations may have experienced genome-wide selective sweeps. This represents the first direct, time-resolved observations of periodic selection in natural populations, a key process predicted by the ecotype model of bacterial diversification.

59 BASIC BIOLOGICAL SCIENCES↗