scholarly journals Distinct human gut microbial taxonomic signatures uncovered with different sample processing and microbial cell disruption methods for metaproteomic analysis

2020 ◽  
Author(s):  
Carmen García-Durán ◽  
Raquel Martínez López ◽  
Inés Zapico ◽  
Enrique Pérez ◽  
Eduardo Romeu ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Stool Sample ◽  
Glass Beads ◽  
Cell Disruption ◽  
Microbial Cell ◽  
Sample Processing ◽  
Human Gut ◽  
Human Gut Microbiota ◽  
Taxonomic Profile ◽  
Stool Samples

AbstractMetaproteomics is as a promising technique for studying the human gut microbiota, because it can reveal the taxonomic profile and also shed light on the functional role of the microbial community. Nevertheless, methods for extracting proteins from stool samples continue to evolve, in the pursuit of optimal protocols for moistening and dispersing the stool sample and for disrupting microbial cells which are two critical steps for ensuring good protein recovery. Here, we evaluated different stool sample processing and microbial cell disruption methods for metaproteomic analyses of human gut microbiota. An unsupervised principal component analysis showed that different methods produced similar human gut microbial taxonomic profiles. An unsupervised two-way hierarchical clustering analysis identified the microbial taxonomic signatures associated with each method. Proteobacteria and Bacteroidetes identification was favored by moistening the stool samples during processing and by disrupting cells with medium-sized glass beads. Ascomycota identification was enhanced by using large-sized glass beads during sample processing for stool dispersion. Euryarchaeota identification was improved with a combination of small and medium-sized glass beads for cell disruption. Assessments of the relative abundance of Firmicutes, Actinobacteria and Spirochaetes improved when ultrasonication was performed before cell disruption with glass beads. The latter method also increased the overall number of identified proteins. Taxonomic and protein functional analyses of metaproteomic data derived from stool samples from six healthy individuals showed common taxonomic profiles. We also detected certain proteins involved in microbial functions relevant to the host and related mostly to particular taxa, such as B12 biosynthesis and short chain fatty acid production carried out mainly by members in the Prevotella genus and the Firmicutes phylum, respectively. Finally, in this metaproteomic study we identified several human proteins, mostly related to the anti-microbial response, which could contribute to determining the beneficial and detrimental relationships between gut microbiota and human cells in particular human diseases.

scholarly journals Distinct Human Gut Microbial Taxonomic Signatures Uncovered With Different Sample Processing and Microbial Cell Disruption Methods for Metaproteomic Analysis

2021 ◽  
Vol 12 ◽  
Author(s):  
Carmen García-Durán ◽  
Raquel Martínez-López ◽  
Inés Zapico ◽  
Enrique Pérez ◽  
Eduardo Romeu ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Stool Sample ◽  
Taxonomic Diversity ◽  
Glass Beads ◽  
Cell Disruption ◽  
Microbial Cell ◽  
Sample Processing ◽  
Human Gut ◽  
Taxonomic Profile ◽  
Stool Samples

The use of metaproteomics for studying the human gut microbiota can shed light on the taxonomic profile and the functional role of the microbial community. Nevertheless, methods for extracting proteins from stool samples continue to evolve, in the pursuit of optimal protocols for moistening and dispersing the stool sample and for disrupting microbial cells, which are two critical steps for ensuring good protein recovery. Here, we evaluated different stool sample processing (SSP) and microbial cell disruption methods (CDMs). The combination of a longer disintegration period of the stool sample in a tube rotator with sonication increased the overall number of identified peptides and proteins. Proteobacteria, Bacteroidetes, Planctomycetes, and Euryarchaeota identification was favored by mechanical cell disruption with glass beads. In contrast, the relative abundance of Firmicutes, Actinobacteria, and Fusobacteria was improved when sonication was performed before bead beating. Tenericutes and Apicomplexa identification was enhanced by moistening the stool samples during processing and by disrupting cells with medium-sized glass beads combined with or without sonication. Human protein identifications were affected by sonication. To test the reproducibility of these gut metaproteomic analyses, we examined samples from six healthy individuals using a protocol that had shown a good taxonomic diversity and identification of proteins from Proteobacteria and humans. We also detected proteins involved in microbial functions relevant to the host and related mostly to specific taxa, such as B12 biosynthesis and short chain fatty acid (SCFA) production carried out mainly by members in the Prevotella genus and the Firmicutes phylum, respectively. The taxonomic and functional profiles obtained with the different protocols described in this work provides the researcher with valuable information when choosing the most adequate protocol for the study of certain pathologies under suspicion of being related to a specific taxon from the gut microbiota.

Sample Processing for Metaproteomic Analysis of Human Gut Microbiota

2021 ◽  
pp. 53-61
Author(s):  
Carmen García-Durán ◽  
Raquel Martínez-López ◽  
Lucía Monteoliva ◽  
Concha Gil
Keyword(s):  
Gut Microbiota ◽  
Sample Processing ◽  
Human Gut ◽  
Human Gut Microbiota

scholarly journals Longitudinal Analysis of Infant Stool Bacteria Communities Before and After Acute Febrile Malaria and Artemether-Lumefantrine Treatment

2018 ◽  
Vol 220 (4) ◽  
pp. 687-698 ◽  
Author(s):  
Rabindra K Mandal ◽  
Rosie J Crane ◽  
James A Berkley ◽  
Wilson Gumbi ◽  
Juliana Wambua ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Bacterial Species ◽  
Enteric Bacteria ◽  
Malaria Episode ◽  
Sequence Variant ◽  
Human Gut ◽  
Human Gut Microbiota ◽  
Significant Difference ◽  
Before And After ◽  
Stool Samples

Abstract Background Gut microbiota were recently shown to impact malaria disease progression and outcome, and prior studies have shown that Plasmodium infections increase the likelihood of enteric bacteria causing systemic infections. Currently, it is not known whether Plasmodium infection impacts human gut microbiota as a prelude to bacteremia or whether antimalarials affect gut microbiota. Our goal was to determine to what degree Plasmodium infections and antimalarial treatment affect human gut microbiota. Methods One hundred Kenyan infants underwent active surveillance for malaria from birth to 10 months of age. Each malaria episode was treated with artemether-lumefantrine (AL). Any other treatments, including antibiotics, were recorded. Stool samples were collected on an approximately biweekly basis. Ten children were selected on the basis of stool samples having been collected before (n = 27) or after (n = 17) a malaria episode and without antibiotics having been administered between collections. These samples were subjected to 16S ribosomal ribonucleic acid gene (V3–V4 region) sequencing. Results Bacterial community network analysis revealed no obvious differences in the before and after malaria/AL samples, which was consistent with no difference in alpha and beta diversity and taxonomic analysis at the family and genus level with one exception. At the sequence variant (SV) level, akin to bacterial species, only 1 of the top 100 SVs was significantly different. In addition, predicted metagenome analysis revealed no significant difference in metagenomic capacity between before and after malaria/AL samples. The number of malaria episodes, 1 versus 2, explained significant variation in gut microbiota composition of the infants. Conclusions In-depth bioinformatics analysis of stool bacteria has revealed for the first time that human malaria episode/AL treatment have minimal effects on gut microbiota in Kenyan infants.

In vitro study of the interaction between human gut microbiota and herbal extracts using willow bark extract as an example

Planta Medica ◽  
2016 ◽  
Vol 81 (S 01) ◽  
pp. S1-S381
Author(s):  
EM Pferschy-Wenzig ◽  
K Koskinen ◽  
C Moissl-Eichinger ◽  
R Bauer
Keyword(s):  
Gut Microbiota ◽  
In Vitro Study ◽  
Vitro Study ◽  
Bark Extract ◽  
Herbal Extracts ◽  
Human Gut ◽  
Human Gut Microbiota ◽  
Willow Bark

Metabolization of the herbal combination STW-5 by human gut microbiota in vitro

2017 ◽  
Author(s):  
EM Pferschy-Wenzig ◽  
A Roßmann ◽  
K Koskinen ◽  
H Abdel-Aziz ◽  
C Moissl-Eichinger ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Human Gut ◽  
Human Gut Microbiota

Substrate use prioritization by a coculture of five species of gut bacteria fed mixtures of arabinoxylan, xyloglucan, β-glucan, and pectin

2020 ◽  
Author(s):  
Y Liu ◽  
AL Heath ◽  
B Galland ◽  
N Rehrer ◽  
L Drummond ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Dietary Fiber ◽  
Plant Cell Wall ◽  
Bacterial Species ◽  
Short Chain ◽  
Emergent Properties ◽  
Human Gut ◽  
Fermentation Products ◽  
Plant Polysaccharides ◽  
Human Gut Microbiota

© 2020 American Society for Microbiology. Dietary fiber provides growth substrates for bacterial species that belong to the colonic microbiota of humans. The microbiota degrades and ferments substrates, producing characteristic short-chain fatty acid profiles. Dietary fiber contains plant cell wall-associated polysaccharides (hemicelluloses and pectins) that are chemically diverse in composition and structure. Thus, depending on plant sources, dietary fiber daily presents the microbiota with mixtures of plant polysaccharides of various types and complexity. We studied the extent and preferential order in which mixtures of plant polysaccharides (arabinoxylan, xyloglucan, β-glucan, and pectin) were utilized by a coculture of five bacterial species (Bacteroides ovatus, Bifidobacterium longum subspecies longum, Megasphaera elsdenii, Ruminococcus gnavus, and Veillonella parvula). These species are members of the human gut microbiota and have the biochemical capacity, collectively, to degrade and ferment the polysaccharides and produce short-chain fatty acids (SCFAs). B. ovatus utilized glycans in the order β-glucan, pectin, xyloglucan, and arabinoxylan, whereas B. longum subsp. longum utilization was in the order arabinoxylan, arabinan, pectin, and β-glucan. Propionate, as a proportion of total SCFAs, was augmented when polysaccharide mixtures contained galactan, resulting in greater succinate production by B. ovatus and conversion of succinate to propionate by V. parvula. Overall, we derived a synthetic ecological community that carries out SCFA production by the common pathways used by bacterial species for this purpose. Systems like this might be used to predict changes to the emergent properties of the gut ecosystem when diet is altered, with the aim of beneficially affecting human physiology. This study addresses the question as to how bacterial species, characteristic of the human gut microbiota, collectively utilize mixtures of plant polysaccharides such as are found in dietary fiber. Five bacterial species with the capacity to degrade polymers and/or produce acidic fermentation products detectable in human feces were used in the experiments. The bacteria showed preferential use of certain polysaccharides over others for growth, and this influenced their fermentation output qualitatively. These kinds of studies are essential in developing concepts of how the gut microbial community shares habitat resources, directly and indirectly, when presented with mixtures of polysaccharides that are found in human diets. The concepts are required in planning dietary interventions that might correct imbalances in the functioning of the human microbiota so as to support measures to reduce metabolic conditions such as obesity.

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