Matthias Hess

Metagenomic discovery of biomass-degrading genes and genomes from cow rumen.

Metagenomic sequencing of biomass-degrading microbes from cow rumen reveals new carbohydrate-active enzymes. The paucity of enzymes that efficiently deconstruct plant polysaccharides represents a major bottleneck for industrial-scale conversion of cellulosic biomass into biofuels. Cow rumen microbes specialize in degradation of cellulosic plant material, but most members of this complex community resist cultivation. To characterize biomass-degrading genes and genomes, we sequenced and analyzed 268 gigabases of metagenomic DNA from microbes adherent to plant fiber incubated in cow rumen. From these data, we identified 27,755 putative carbohydrate-active genes and expressed 90 candidate proteins, of which 57% were enzymatically active against cellulosic substrates. We also assembled 15 uncultured microbial genomes, which were validated by complementary methods including single-cell genome sequencing. These data sets provide a substantially expanded catalog of genes and genomes participating in the deconstruction of cellulosic biomass.

The microbial nitrogen cycling potential is impacted by polyaromatic hydrocarbon pollution of marine sediments

During hydrocarbon exposure, the composition and functional dynamics of marine microbial communities are altered, favoring bacteria that can utilize this rich carbon source. Initial exposure of high levels of hydrocarbons in aerobic surface sediments can enrich growth of heterotrophic microorganisms having hydrocarbon degradation capacity. As a result, there can be a localized reduction in oxygen potential within the surface layer of marine sediments causing anaerobic zones. We hypothesized that increasing exposure to elevated hydrocarbon concentrations would positively correlate with an increase in denitrification processes and the net accumulation of dinitrogen. This hypothesis was tested by comparing the relative abundance of genes associated with nitrogen metabolism and nitrogen cycling identified in 6 metagenomes from sediments contaminated by polyaromatic hydrocarbons from the Deepwater Horizon (DWH) oil spill in the Gulf of Mexico, and 3 metagenomes from sediments associated with natural oil seeps in the Santa Barbara Channel. An additional 8 metagenomes from uncontaminated sediments from the Gulf of Mexico were analyzed for comparison. We predicted relative changes in metabolite turnover as a function of the differential microbial gene abundances, which showed predicted accumulation of metabolites associated with denitrification processes, including anammox, in the contaminated samples compared to uncontaminated sediments, with the magnitude of this change being positively correlated to the hydrocarbon concentration and exposure duration. These data highlight the potential impact of hydrocarbon inputs on N cycling processes in marine sediments and provide information relevant for system scale models of nitrogen metabolism in affected ecosystems.

Thermoacidophilic proteins for biofuel production

Growing concerns about global climate change and energy dependence have led to an increased effort to reduce carbon emissions. A considerable reduction could be achieved by using biofuels from lignocellulosic biomass instead of fossil fuels. One major bottleneck of biofuel production from lignocellulose is the availability of efficient and inexpensive biocatalysts (i.e. alcohol dehydrogenases, cellulases and esterases) that are active and stable at high temperatures and low pH values. Although heterologous gene expression is used effectively to obtain recombinant proteins derived from mesophiles, the production of thermoacidophilic proteins is often unsuccessful. Some of the reasons for this failure and potential solutions for an increased production of novel extremophilic biocatalysts are discussed here.

Temporal dynamics of fibrolytic and methanogenic rumen microorganisms during in situ incubation of switchgrass determined by 16s rRNA gene profiling

The rumen microbial ecosystem is known for its biomass-degrading and methane-producing phenotype. Fermentation of recalcitrant plant material, comprised of a multitude of interwoven fibers, necessitates the synergistic activity of diverse microbial taxonomic groups that inhabit the anaerobic rumen ecosystem. Although interspecies hydrogen (H2) transfer, a process during which bacterially generated H2 is transferred to methanogenic Archaea, has obtained significant attention over the last decades, the temporal variation of the different taxa involved in in situ biomass-degradation, H2 transfer and the methanogenesis process remains to be established. Here we investigated the temporal succession of microbial taxa and its effect on fiber composition during rumen incubation using 16S rRNA amplicon sequencing. Switchgrass filled nylon bags were placed in the rumen of a cannulated cow and collected at nine time points for DNA extraction and 16S pyrotag profiling. The microbial community colonizing the air-dried and non-incubated (0 h) switchgrass was dominated by members of the Bacilli (recruiting 63% of the pyrotag reads). During in situ incubation of the switchgrass, two major shifts in the community composition were observed: Bacilli were replaced within 30 min by members belonging to the Bacteroidia and Clostridia, which recruited 34 and 25% of the 16S rRNA reads generated, respectively. A second significant shift was observed after 16 h of rumen incubation, when members of the Spirochaetes and Fibrobacteria classes became more abundant in the fiber-adherent community. During the first 30 min of rumen incubation ~13% of the switchgrass dry matter was degraded, whereas little biomass degradation appeared to have occurred between 30 min and 4 h after the switchgrass was placed in the rumen. Interestingly, methanogenic members of the Euryarchaeota (i.e., Methanobacteria) increased up to 3-fold during this period of reduced biomass-degradation, with peak abundance just before rates of dry matter degradation increased again. We hypothesize that during this period microbial-mediated fibrolysis was temporarily inhibited until H2 was metabolized into CH4 by methanogens. Collectively, our results demonstrate the importance of inter-species interactions for the biomass-degrading and methane-producing phenotype of the rumen microbiome—both microbially facilitated processes with global significance.

High‐Throughput In Vitro Glycoside Hydrolase (HIGH) Screening for Enzyme Discovery

A high-throughput protein-expression and screening method (HIGH method, see picture) provides a rapid approach to the discovery of active glycoside hydrolases in environmental samples. Finally, HIGH screening combines cloning, protein expression, and enzyme hydrolysis in one pot; thus, the entire process from gene expression to activity detection requires only three hours.

Identification of novel biomass-degrading enzymes from genomic dark matter: Populating genomic sequence space with functional annotation

Although recent nucleotide sequencing technologies have significantly enhanced our understanding of microbial genomes, the function of ∼35% of genes identified in a genome currently remains unknown. To improve the understanding of microbial genomes and consequently of microbial processes it will be crucial to assign a function to this “genomic dark matter.” Due to the urgent need for additional carbohydrate‐active enzymes for improved production of transportation fuels from lignocellulosic biomass, we screened the genomes of more than 5,500 microorganisms for hypothetical proteins that are located in the proximity of already known cellulases. We identified, synthesized and expressed a total of 17 putative cellulase genes with insufficient sequence similarity to currently known cellulases to be identified as such using traditional sequence annotation techniques that rely on significant sequence similarity. The recombinant proteins of the newly identified putative cellulases were subjected to enzymatic activity assays to verify their hydrolytic activity towards cellulose and lignocellulosic biomass. Eleven (65%) of the tested enzymes had significant activity towards at least one of the substrates. This high success rate highlights that a gene context‐based approach can be used to assign function to genes that are otherwise categorized as “genomic dark matter” and to identify biomass‐degrading enzymes that have little sequence similarity to already known cellulases. The ability to assign function to genes that have no related sequence representatives with functional annotation will be important to enhance our understanding of microbial processes and to identify microbial proteins for a wide range of applications. Biotechnol. Bioeng. 2014;111: 1550–1565.

Metagenomic analysis of microbial consortium from natural crude oil that seeps into the marine ecosystem offshore Southern California.

Crude oils can be major contaminants of the marine ecosystem and microorganisms play a significant role in the degradation of its main constituents. To increase our understanding of the microbial hydrocarbon degradation process in the marine ecosystem, we collected crude oil from an active seep area located in the Santa Barbara Channel (SBC) and generated a total of about 52 Gb of raw metagenomic sequence data. The assembled data comprised ∼500 Mb, representing ∼1.1 million genes derived primarily from chemolithoautotrophic bacteria. Members of Oceanospirillales, a bacterial order belonging to the Deltaproteobacteria, recruited less than 2% of the assembled genes within the SBC metagenome. In contrast, the microbial community associated with the oil plume that developed in the aftermath of the Deepwater Horizon (DWH) blowout in 2010, was dominated by Oceanospirillales, which comprised more than 60% of the metagenomic data generated from the DWH oil plume. This suggests that Oceanospirillales might play a less significant role in the microbially mediated hydrocarbon conversion within the SBC seep oil compared to the DWH plume oil. We hypothesize that this difference results from the SBC oil seep being mostly anaerobic, while the DWH oil plume is aerobic. Within the Archaea, the phylum Euryarchaeota, recruited more than 95% of the assembled archaeal sequences from the SBC oil seep metagenome, with more than 50% of the sequences assigned to members of the orders Methanomicrobiales and Methanosarcinales. These orders contain organisms capable of anaerobic methanogenesis and methane oxidation (AOM) and we hypothesize that these orders - and their metabolic capabilities - may be fundamental to the ecology of the SBC oil seep.

Metagenomes from two microbial consortia associated with Santa Barbara seep oil

The metagenomes from two microbial consortia associated with natural oils seeping into the Pacific Ocean offshore the coast of Santa Barbara (California, USA) were determined to complement already existing metagenomes generated from microbial communities associated with hydrocarbons that pollute the marine ecosystem. This genomics resource article is the first of two publications reporting a total of four new metagenomes from oils that seep into the Santa Barbara Channel.

Metagenomic Sequencing of Two Salton Sea Microbiomes

ABSTRACT The Salton Sea is the largest inland body of water in California, with salinities ranging from brackish freshwater to hypersaline. The lake experiences high nutrient input, and its surface water is exposed to temperatures up to 40°C. Here, we report the community profiles associated with surface water from the Salton Sea.

Metagenome sequencing of the prokaryotic microbiota of the hypersaline and meromictic soap lake, washington.

ABSTRACT Soap Lake is a small saline lake in central eastern Washington that is sharply stratified into two layers. In addition to being highly alkaline (~pH 10), Soap Lake also contains high concentrations of sulfide. Here, we report the community profile of the prokaryotic microbiota associated with Soap Lake surface water.