This application seeks funding to purchase an Illumina MiSeq system. This instrument is a cutting- edge, next generation benchtop DNA sequencer that will support studies of the gut microbiome by our VA funded investigators. The MiSeq system includes integrated software for automated generation of DNA clonal clusters by bridge amplification, sequencing, primary and secondary analysis. The system includes an embedded touchscreen monitor and on-instrument computer, dual surface imaging capability, MiSeq Software Suite, installation kits and standards, installation and training, and 12 months warranty (including parts and labor). The MiSeq system is capable of generating up to 15 Gb per run using Nextera kits in combination with Illumina's validated TruSeq chemistry. The instrument footprint requires less than 2 square feet of benchtop space and is a stand-alone instrument that does not need dedicated ancillary amplification or computing/IT input. Metagenomic surveys of microbial populations are performed using the prokaryotic 16S ribosomal RNA (rRNA) gene, which contains conserved and variable regions that facilitate sequencing and phylogenetic classification of the gut microbiome. Using the MiSeq system, 16S metagenomics studies can achieve bacterial species-level identification efficiently. The MiSeq system will allow expansion of ongoing and currently funded research on the following projects of our 5 major users: 1) The Kuhn lab is studying the role of serotonin in the non-motor symptoms of Parkinson's disease and it was recently demonstrated that the gut microbiome plays a very critical role in the development of this neurodegenerative disorder. Studies of the gut microbiome in a mouse model that lacks brain serotonin (developed in the Kuhn lab for these studies) will add a new dimension to this project; 2) the Majumdar lab is studying colorectal cancer and how alterations in the gut microbiome can define racial disparities in this malignancy. Dr. Majumdar has carried out limited sequencing of the human gut microbiome already and the MiSeq system will allow his lab to expand these interesting studies in Veterans with colorectal cancer; 3) the Kowluru lab is studying diabetes and the islet cells function. In light of emerging work that has demonstrated that the pathological path followed by diabetes is significantly influenced by the gut microbiome, Dr. Kowluru will use the MiSeq to study how currently used therapeutics for diabetes changes the gut microbiome and improves outcomes: 4) The Mateika lab is studying the role of serotonin in respiratory function in spinal cord injury. A single published paper has recently suggested that spinal cord injury causes a significant alteration in the gut microbiome and Dr. Mateika will study how respiratory and sleep alterations associated with spinal cord injury are modulated by the gut microbiome; and 5) the Rishi lab is studying new therapeutics for the treatment of malignancies and a very recent publication revealed that cancer cell resistance to selected chemotherapeutics is mediated by specific bacteria in the gut microbiome. Dr. Rishi's research is focusing on cancer cell multidrug resistance using in vivo models. With the MiSeq system, the Rishi lab will be able to expand its approach to the study of cancer treatment. Together, these five investigators study some of the most prevalent clinical conditions affecting Veterans (i.e., Parkinson's disease, diabetes/obesity, cancer and spinal cord injury). Emerging studies are showing clearly that these same diseases are significantly modulated by the gut microbiome. The ability of our investigators to carry out 16S rRNA sequencing in their studies will not only add new and innovative dimensions to the individual projects, it will also enhance our ability to translate these findings to the clinic and improve the health care delivered to our Veterans.
The broad, long-term objectives of this ShEEP application for funding to purchase a MiSeq system are to initiate studies of the gut microbiome in our ongoing projects on Parkinson's disease, diabetes/obesity, cancer and spinal cord injury and to leverage existing VA funding to expand these projects through the study of the gut microbiome. The MiSeq system is a state-of-the-art sequencer that will help move our funded investigators into new and innovative research realms - the gut microbiome and the gut-brain axis. The gut microbiome regulates numerous physiological systems in health and when it becomes unbalanced, numerous pathological conditions can emerge. The MiSeq will therefore add an important dimension to the research of numerous VA funded investigators at our station and enhance our ability to collaborate on the emerging gut-brain axis.
