Project description:The aim of this study was to develop a suitable method to preserve fecal samples for metaproteomics analyses when flash-freezing is not an option. Fecal samples were collected from conventional adult C57BL/6 mice and combined into a fecal master mix. The fecal master mix was then split into 48 subsamples that were subjected to different preservation treatments. The following six preservation methods were tested: flash-freezing in liquid nitrogen followed by storage at -80°C, immersion in RNAlater® and storage at room temperature, immersion in RNAlater® and immediate storage at -80°C, immersion in 95% ethanol and storage at room temperature, immersion in a RNAlater-like buffer “NAP buffer” and storage at room temperature, and immersion in an autoclaved RNAlater-like buffer “Autoclaved NAP buffer” and storage at room temperature. Proteins were extracted from the samples after being stored for 1 and 4 weeks. There were 4 replicates per treatment and time-point. Samples were analyzed by LC-MS/MS and the data were analyzed with Proteome Discoverer against a large database of mouse microbiota protein sequences.
Project description:Sample preservation method that maintains transcripts in viable single cells and so allows to disconnect time and place of sampling from subsequent processing steps.
2017-02-13 | GSE85534 | GEO
Project description:Effects of preservation and storage conditions on the fecal microbiome
| PRJEB13595 | ENA
Project description:Further Effects of preservation and storage conditions on the fecal microbiome
Project description:Bacterial persistence, found in dormant and starved cells, is a health threat due to transient antibiotic tolerance. Harnessing a novel method for persister generation, we determined the proteome, metabolite levels and the physiology of E.coli persisters in and during entry into dormancy and starvation. In contrast to starved persisters, dormant persisters present in nutrient-rich conditions produced energy and grew, while both types had extremely low metabolite pools. The proteome of dormant cells governed by starvation response reached a unique state characterized by diminished anabolism, stress response and preservation of central metabolism protein levels. While starved cells approaches the same proteome, the limited carbon and energy source did not allow them to reach it, which caused their higher sensitivity to certain antibiotics. We present a conceptual model in which depleted metabolite pools resulting from initial persistence triggers provide a primitive, feed-forward starvation signal that sustains the growing persistent phenotype.
Project description:Analysis of breast cancer survivors' gut microbiota after lifestyle intervention, during the COVID-19 lockdown, by 16S sequencing of fecal samples.
Project description:Bone collagen is an important organic material for isotopic measurement, radiocarbon and paleoproteomic analyzes, to provide information on diet, dating, taxonomic identification. Current paleoproteomics methods are destructive and require from a few milligrams to several tenths of milligrams of bone for analysis. In many cultures, bones are raw materials for artefact which are conserved in museum which hampers to damage these precious objects during sampling. Here, we describe a minimal sampling method that identifies collagen, taxonomy and post-translational modifications from Holocene and Upper Pleistocene bones dated to 130,000 and 5,000 years ago using dermatological skin tape-discs for sampling. The sampled bone micro-powder were digested following our highly optimized eFASP protocol, then analyzed by MALDI FTICR MS and LC-MS/MS for identifying the genus taxa of the bones. We show that this low-invasive sampling does not deteriorate the bones and achieves results similar to those obtained by destructive sampling. Moreover, this sampling method can be performed at archaeological sites or in museums.
Project description:Gut microbiome modulates the host immune development, yet the functional contribution of gut fungi remains elusive. We previously showed that mice colonized only with fungi displayed allergic features and fecal metabolite profiles similar to germ-free mice. To gain insights into the functional changes attributed to fungal colonization, we performed proteomic analyses of feces and small intestine of gnotobiotic mice colonized with either bacteria, fungi, or both. Comparison of fecal metaproteomic profiles between mouse groups yielded broad changes in the relative levels of bacterial, fungal and mouse proteins. Many of the detected fungal proteins have been previously reported as a part of extracellular vesicles and having immunomodulating properties. Changes in the levels of mouse proteins derived from the small intestine impacted essential cellular pathways, including lipid metabolism and apoptosis. The results show how fungal colonization impacts the host proteome and suggest an influence on the host final cellular phenotype.