Dunham, S. J. B. | bioRxiv Science

bioRxiv2022

Longitudinal analysis of the gut microbiome in the 5xfAD mouse model of Alzheimer's disease

INTRODUCTIONMicrobial exposures impact Alzheimers disease (AD), and a baseline understanding of AD model microbiomes is vital for improving model efficacy. We describe the evaluation of the microbiome and metabolome in longitudinal samples from the 5xfAD transgenic mouse model. METHODSCecal and fecal samples from 5xfAD and wild-type B6J (WT) animals from 4- 18 months of age were subjected to DNA extraction and shotgun Illumina sequencing. Metabolomics was performed on plasma and feces from a subset of animals. RESULTSSignificant sex, age, and cage-specific differences were observed in the microbiome. Eight bacteria species were elevated in older 5xfAD mice and nine were depleted, including Turicibacter spp. Contradicting published findings covering persons with AD, plasma measurements revealed elevated serotonin in 18 month 5xfAD animals. DISCUSSIONTaken together, these findings strengthen the link between Turicibacter abundance and AD and provide a basis for further microbiome studies of murine models for AD.

microbiology↗

bioRxiv2021

Capturing actively produced microbial volatile organic compounds from human associated samples with vacuum assisted sorbent extraction

Volatile organic compounds (VOC) from biological samples have unknown origins. VOCs may originate from the host or different microbial organisms from a microbial community. In order to disentangle the origin of microbial VOCs, we performed volatile headspace analysis of bacterial mono- and co-cultures of Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii, and stable isotope probing in biological samples of feces, saliva, sewage, and sputum. We utilized mono- and co-cultures to identify volatile production from individual bacterial species or in combination with stable isotope probing to identify the active metabolism of microbes from the biological samples. To extract the VOCs, we employed vacuum assisted sorbent extraction (VASE). VASE is an easy-to-use commercialized solvent-free headspace extraction method for semi-volatile and volatile compounds. The lack of solvents and the near vacuum conditions used make developing a method relatively easy and fast when compared to other extraction options like tert-butylation and solid phase microextraction. However, VASE does not work on nonvolatile compounds, thus excluding many protein analyses and heavy stable isotope labelling experiments. Using the workflow described here, we identified volatile signatures from mono- and co-cultures indicating there were volatiles specific to certain microbes or co-cultures. Furthermore, analysis of the stable isotope probing of biological samples identified VOCs that were either commonly or uniquely produced from the different human derived biological samples. Here we present the general workflow and experimental considerations of VASE. SUMMARYFrom this protocol, readers will be able to extract volatile organic compounds from a biological sample with the vacuum assisted sorbent extraction method, run samples on a GC-MS with the use of the Entech autosampler, and analyze data.

biochemistry↗