Dataset Information

The proteome of extracellular vesicles secreted in vitro by porcine microbiota grown in complex β-mannan

ABSTRACT: The secretion of Extracellular Vesicles ((EVs) is a common process in both prokaryotic and eukaryotic cells for intercellular communication, survival, and pathogenesis. Previous studies have illustrated the presence of EVs in supernatants from pure cultures of bacteria, including Gram-positive and Gram-negative glycan-degrading gut commensals. However, the composition of vesicles formed in complex microbial communities has not been clearly defined. In a recent paper, we reported that wood-derived complex β-mannan, which shares structural similarity with conventional dietary fibers, can be used to modulate the porcine gut microbiota composition and activity. In this paper, we investigated the production, size, composition, and proteome of EVs secreted by the pig fecal microbiota after 24 h enrichment on complex β-mannan. Using transmission electron microscopy and nanoparticle tracking analysis, we identified EVs with an average size of 105 nm. We utilized mass spectrometry-based metaproteomic profiling of EV proteins against a database of 355 metagenome-assembled genomes (MAGs) from the porcine colon and thereby identified 303 proteins, of which 55% were only detected in the EVs isolated from the β-mannan culture. Most of the proteins extracted from the EVs isolated from the culture grown on β-mannan mapped to two MAGs, MAG53 and MAG272, belonging to the orders Clostridiales and Bacilli, respectively. Furthermore, the MAG with the third-most-detected proteins was MAG 343 belonging to the order Enterobacteriales. The most abundant proteins detected in the β-mannan proteome were involved in translation, energy production, amino acid and carbohydrate transport, as well as metabolism. Overall, our results demonstrate the release of EVs in a complex microbial community whereby the protein content of the EVs reflects the response of specific microbes to the available carbohydrate source.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Sus Scrofa Domesticus (domestic Pig)

SUBMITTER: Leidy Lagos

LAB HEAD: Leidy Lagos

PROVIDER: PXD017420 | Pride | 2021-09-09

REPOSITORIES: Pride

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Isolation and Characterization of Extracellular Vesicles Secreted In Vitro by Porcine Microbiota.

Lagos Leidy L Leanti La Rosa Sabina S Ø Arntzen Magnus M Ånestad Ragnhild R Terrapon Nicolas N Gaby John Christian JC Westereng Bjørge B

Microorganisms 20200630 7

The secretion of extracellular vesicles, EVs, is a common process in both prokaryotic and eukaryotic cells for intercellular communication, survival, and pathogenesis. Previous studies have illustrated the presence of EVs in supernatants from pure cultures of bacteria, including Gram-positive and Gram-negative glycan-degrading gut commensals. However, the isolation and characterization of EVs secreted by a complex microbial community have not been clearly reported. In a recent paper, we showed t ...[more]

PMID: 32630095

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Project description:Current developments have led to a reconsidering of energy policy in many countries with the aim of increasing the share of renewable energies in the energy supply, where the anaerobic digestion (AD) of biomass to produce methane also plays an important role. To improve biomass digestion while ensuring overall process stability, microbiome-based management strategies became more important. By applying combined metagenome and metaproteome, as well as metagenomically assembled genome (MAG)-centric analyses, it is possible to determine not only the functional potential but also the expressed functions of the entire microbial community and also individual MAGs. This approach was used in this study for the production-scale biogas plant 35 (BP35) consisting of three digesters which were operated differently regarding process temperatures, feedstocks and other process parameters. Different process conditions were hypothesized to result in specific microbiome adaptations and differentially abundant metabolic functions in the digesters. Based on metagenomic single-read analyses, several taxa residing in the three digesters of BP35 were shown to correlate with the corresponding substrates and temperatures. In particular, the genus Defluviitoga showed the strongest correlation to the process temperature and the genus Acetomicrobium featured a direct correlation to the concentartions of different acids including acetic acid. Analysis of the functional potential and expressed functions of the entire microbial community of the three digesters revealed that the genes and key enzymes relevant for the biogas process were present and also expressed. Differences between the abundances of certain genes and expressed enzymes could be related to the specific parameters of the corresponding digesters. Regarding the biogas related metabolic pathways, MAG-centric metagenomics and metaproteomics indicated the functional potential and the actual expressed metabolic functions of certain MAGs that are differentially abundant in the three digesters. These MAGs, belonging to the phylum Firmicutes, the class Bacilli and the orders Caldicoprobacterales and Bacteroidales showed a specific metabolic activity within the three digesters and have important roles in the hydrolysis, acidogenesis or acetogenesis of the anaerobic digestion process. An archaeal MAG assigned to the species Methanothermobacter wolfeii was the most abundant and highly active hydrogenotrophic methanogen in digester 3 featuring an operation temperature of 54 °C. Beside the MAGs that were differentially abundant in the three digesters, also MAGs which were more evenly distributed were analyzed. The most abundant and highly active MAG in all digesters belongs to the class Limnochordia and was shown to be ubiquitous in all three digesters and exhibit activity in a variety of pathways representing hydrolysis as well as the acido- and acetogenesis steps of the biogas process. Other MAGs assigned to the phylum Firmicutes, genus Acetomicrobium and the hydrogenotrophic species Methanoculleus thermohydrogenotrophicum were also shown to be more evenly distributed and active in the three digesters. Corresponding taxa appeared to be more resilient to the different process parameters of the three digesters, and therefore, may support a stable biogas process. Overall, the combined metagenome and metaproteome analysis of biogas digesters helps to gain deeper insights into the composition of the whole microbial community, biogas related pathways and their expression, which could contribute to an improved microbiome-based process management in the future.

			Action	DRS
	180112_Leidly_01.raw	Raw
	180112_Leidly_02.raw	Raw
	180112_Leidly_03.raw	Raw
	180112_Leidly_04.raw	Raw
	190218LeidyEVbasedonMQ14.sps	Other