Enter the text here that is the new abstr Recent advances in large-scale DNA sequencing have the potential for rapid advances in correlating disease phenotypes with genotypes. Infectious disease research has long focused on the pathogen's role in infection but less so on the impact of the host genotype. The current epidemic of Methicillin Resistant Staphylococcus aureus (MRSA) infection is a Grand Opportunity to apply new genetic methods to understanding this disease. There are multiple outcomes from MRSA infection, ranging from curable localized infection to devastating invasive infection. The project will study 500 subjects with MRSA infections by applying high throughput DNA sequencing to candidate genes involved in immune and other host responses to infection to correlate the outcome of infection with the host genotype. In addition, MRSA isolates from these same patients will be host immune responses and the arsenal of bacterial virulence factors on outcomes of community acquisition of MRSA is largely unknown. Our long-range objective is to seek a biological explanation for the wide spectrum of disease manifestations caused by CA-MRSA with the immediate goal of understanding the presentation of severe invasive infections in otherwise healthy individuals. We hypothesize that the outcome of CA-MRSA acquisition is determined by interactions between the genetic composition of the infecting bacterium and polymorphisms in relevant host genes involved in immune, inflammatory, and other host defense responses. 1. Development of a capture system for host defense genes. a. Selection of candidate genes and oligonucleotide probes;b. Development of solution capture strategy: WUCap or other state-of-the-art methods will be employed;c. Validation of the method with PCR-resequencing of a subset of genes;use of VarScan for data analysis. 2. Recruitment, phenotyping, and sampling of 500 subjects (infected and controls). a. Recruitment from up to 6 different clinical sites: i. Washington University School of Medicine will provide DNA samples from 200 subjects and 450 bacterial isolates from these subjects;ii. Texas Childrens Hospital (Houston) will provide DNA samples from 150 subjects and, if needed, additional bacterial isolates from these subjects;iii. The University of Chicago, will provide DNA samples from 150 subjects and, if needed, additional bacterial isolates from these subjects. b. Phenotyping of the clinical presentation of each infection;c. Sampling: isolation of human and bacterial DNA from these subjects. 3. Sequencing of host defense genes in 500 subjects using the capture methodology in Aim 1 and next generation sequencing such as Illumina or 454. 4. Bioinformatic analysis of sequences for candidate mutations in host defense genes using the Genome Center at Washington University variant detection pipeline, currently in use for analysis of whole genome sequences. a. SNP detection;b. Small indel detection;c. Prediction of mutations with functional consequences. 5. Validation of mutations using alternative sequencing methods such as PCR-based Sanger resequencing. 6. Sequencing of bacterial genomes using the Illumina and 454 platforms. 7. Analysis of bacterial sequences for variants using the Genome Center at Washington University variant detection pipeline as modified for bacterial genome analysis. 8. Validation of candidate bacterial variants using consensus data from at least two different next generation sequencing platforms or alternative sequencing methods such as PCR-based Sanger resequencing. 9. Correlation of host and bacterial variants with clinical phenotypes and assessment of statistical confidence in the correlations.
The current epidemic of Methicillin Resistant Staphylococcus aureus (MRSA) infection is a Grand Opportunity to apply new genetic methods to understanding this disease. There are multiple outcomes from MRSA infection, ranging from curable localized infection to devastating invasive infection. The project will focus on the cause of the severe invasive infection, allowing rapid prediction of infection outcome and ultimately leading to rational therapeutic intervention.
| Rodriguez, Marcela; Hogan, Patrick G; Satola, Sarah W et al. (2015) Discriminatory Indices of Typing Methods for Epidemiologic Analysis of Contemporary Staphylococcus aureus Strains. Medicine (Baltimore) 94:e1534 |
| Fritz, Stephanie A; Tiemann, Kristin M; Hogan, Patrick G et al. (2013) A serologic correlate of protective immunity against community-onset Staphylococcus aureus infection. Clin Infect Dis 56:1554-61 |
