Invasive fungal infections are currently important causes of morbidity and mortality among immunocompromised patients. Because the clinical presentation of mycoses is nonspecific, diagnosis is often delayed or erroneous with currently available assays. The present proposal addresses the diagnosis of infections with the major fungal killer, Aspergillus fumigatus, although success with this approach could also be extended to methods to diagnose other microbial opportunistic infections. This study proposes to develop a novel methodology, magnetic-based nanoparticle array assay, which has the potential for very rapid diagnostic capabilities, great sensitivity, specificity, and multiplex capabilities that can extend eventually to other pathogens. The nanoparticle array detects small perturbations in magnetic fields reflected by changes in electrical resistance, such as can occur, for example, with hybridization of a known fragment of fungal DNA adherent to the nanoarray to circulating amounts of that DNA present in body fluids of infected patients. This is the approach of the present project, which will first select the most suitable Aspergillus DNA target to be detected by the nanoarray. Then the reproducibility, sensitivity and specificity of the array with known quantities of target DNA will be assessed in vitro. With suitable characteristics of the assay developed, mouse models of Aspergillus infection (which closely mimic various types of human aspergillosis) will be refined to provide samples of body fluids under controlled replicate conditions, allowing both concurrent quantitation of the amount of infection present and certainty of the time frame of the onset of infection. In the second phase of the studies, the possibility of extension of the approach to other, similar infections will be explored. Another novel technology, RNA-Seq, will use ultra high-throughput sequencing to delineate which genes are most abundantly expressed during infection, and sequence them, with the idea of generating new DNA or RNA targets that might be utilized in the array to give it greater specificity and sensitivity. Finally, the nanoarray platform will be assessed, in a pilot study, for its ability to use microbial (Aspergillus, as prototype) proteins as an alternative target detectable by the use of antibody;RNA and protein targets have the advantage of amplification at the site of infection, possibly facilitating detection at accessible sampling sites, such as blood and other body fluids.