Developing a Method to Study the Impact of Microbial DNA on the Brain Related to Alzheimer’s Disease – Can Food Make a Change?

Abstract: Understanding the gut-brain-microbiota axis can help us provide a new tool to manage conditions such as Alzheimer’s. However, a major challenge is selectively amplifying the bacterial DNA which is present at a much lower level than the host DNA. In this thesis, we aim to develop a methodology to extract, purify and quantify the bacterial DNA in the brain tissue of mice. The methodology that can be used for further application studying gut-brain-microbiota axis, which in this case is comparison of the microbial DNA composition in mice with Alzheimer’s disease vs. healthy littermates fed with natural foods. A major hurdle to this approach is to identify the possible bacterial contamination during the extraction step and to eliminate the host DNA. Brain is a unique and challenging organ to study. The amount of lipids content is second abundance to adipose tissue among others and the solid blood-brain-barrier (BBB) delicately protects the central nerve system (CNS) from microbe’s invasion. The extreme low amount of microbial DNA compared to host DNA after the extraction complicate the efficiency to characterize the microbiota community in the brain tissue. Therefore, in this study we evaluated the bacterial DNA extraction efficiency from murine brain samples and a human saliva, using two commercially available kits (Qiagen DNeasy Blood & Tissue Kits and Promega Wizard® Genomic DNA Purification Kit). While the host DNA depletion was performed using NEBNext Microbiome DNA Enrichment kit. The purified and enriched DNA was further analyzed using gel electrophoresis and finally sequenced by next-generation sequencing targeting the V4 region of the 16S ribosomal RNA gene. The percentage of microbiome DNA component increased in Qiagen compared to Promega method with host DNA depletion. DNA extraction with host DNA depletion influences microbial community composition underlying the need for careful selection of DNA extraction kit and usage of microbiome DNA enrichment to improve recovery from a range of bacterial taxa. Overall, the bacterial community profile of brain tissue was not well characterized in this study which needed further investigation.