Project description:This study aimed to analyze changes in gut microbiota composition in mice after transplantation of fecal microbiota (FMT, N = 6) from the feces of NSCLC patients by analyzing fecal content using 16S rRNA sequencing, 10 days after transplantation. Specific-pathogen-free (SPF) mice were used for each experiments (N=4) as controls.
Project description:Leaf-cutting ants of the genera Acromyrmex and Atta live in mutualistic symbiosis with a basidiomycete fungus (Leucocoprinus gongylophorus), which they cultivate as fungal gardens in underground nest chambers. The ants provide the fungus with a growth substrate consisting of freshly cut leaf fragments. After new leaf fragments are brought into the nest, the ants chew them into smaller pieces and apply droplets of fecal fluid to the leaf pulp before depositing this mixed substrate in the fungus garden and inoculating it with small tufts of mycelium from older parts of the garden. Previous work has shown that the fecal fluid contains a range of digestive enzymes including proteases, amylases, chitinases, cellulases, pectinases, hemicellulases and laccases, and that most of these enzymes are produced by the fungal symbiont in specialized structures called gongylidia that the ants eat. After ingestion, the enzymes apparently pass unharmed through the alimentary channel of the ants and end up in the fecal fluid. Most likely this complex system is an adaptation of the ant-fungus symbiosis to a herbivorous lifestyle, as the ancient ancestors of the ants and the fungus lived as hunter-gatherers and saprotrophs, respectively. The promise of fecal fluid for getting insight into the molecular adaptations that enables the ant-fungus holosymbiont to live as a herbivore, led us to investigate the fecal fluid proteome using LC-MS/MS in order to get a more comprehensive picture of the repertoire of proteins present.
Project description:We report the sequencing of bacterial species across four environments in which C9orf72 loss of function mice were studied as well as mice that received fecal transplantation. Our study elucidates bacterial communities that associate with pro-inflammatory or pro-survival outcomes in this model of ALS/FTD with features of autoimmunity and systemic and neural inflammation.
Project description:<p><strong>BACKGROUND:</strong> The human intestinal microbiome plays a central role in overall health status, especially in early life stages. 16S rRNA amplicon sequencing is used to profile its taxonomic composition; however, multiomic approaches have been proposed as the most accurate methods for study of the complexity of the gut microbiota. In this study, we propose an optimized method for bacterial diversity analysis that we validated and complemented with metabolomics by analyzing fecal samples.</p><p><strong>METHODS:</strong> Forty-eight different analytical combinations regarding (1) 16S rRNA variable region sequencing, (2) a feature selection approach, and (3) taxonomy assignment methods were tested. A total of 18 infant fecal samples grouped depending on the type of feeding were analyzed by the proposed 16S rRNA workflow and by metabolomic analysis.</p><p><strong>RESULTS:</strong> The results showed that the sole use of V4 region sequencing with ASV identification and VSEARCH for taxonomy assignment produced the most accurate results. The application of this workflow showed clear differences between fecal samples according to the type of feeding, which correlated with changes in the fecal metabolic profile.</p><p><strong>CONCLUSION:</strong> A multiomic approach using real fecal samples from 18 infants with different types of feeding demonstrated the effectiveness of the proposed 16S rRNA-amplicon sequencing workflow.</p>
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.
2021-11-25 | PXD024115 | Pride
Project description:16S rRNA amplicon sequencing of human fecal samples
Project description:A rapid ex vivo microbiome assay and metaproteomics approach was used for rapid evaluation of the cultivability of bio-banked live microbiota, which shows minimal detrimental influences of long-term freezing in deoxygenated glycerol buffer on the cultivability of fecal microbiota.