The increasing emergence of bacterial strains resistant to antibiotics is far outpacing our ability to develop novel therapies to treat these infections. Further, the methodologies used to both diagnose infection and predict appropriate courses of treatment are antiquated. Thus, novel ways of thinking about diagnosing and treating infectious diseases must be developed. By merging the powerful fields of chemical biology and bacterial genetics/genomics, we hope to provide insight into possible new paradigms for addressing infectious diseases. The goal of the Hung lab is three fold: 1) to understand mechanisms of bacterial pathogenesis that might better predict both host and pathogen genes that are required for survival of the pathogen in vivo and that might ultimately suggest new ways to intervene based on disrupting the pathogen-host interaction; 2) identifying bacterial functions essential for in vivo survival and persistence (survival in the face of antibiotic therapy), and small molecule tools with which to probe the biology of these functions in vivo; and 3) identifying transcriptional responses to antibiotic stress and developing new tools to both rapidly diagnose a pathogen and determine antibiotic susceptibility based on these responses. Examples of approaches taken include high-throughput forward chemical screens, genomic characterization including single cell analysis, classical forward and reverse genetics, and multiple in vivo models to address these areas, which are described in more detail below.

1) Host and pathogen genetic requirements for bacterial survival in vivo. Using models of Mycobaterium tuberculosis, Pseudomonas aeruginosa, Salmonella typhimurium, and Klebsiella pneumonia, we are conducting classical forward and reverse genetic and chemical genetic screens coupled with high content imaging to identify bacterial functions that are required for survival within host cells. To explore host factors important for bacterial pathogenesis, we are using a combination of chemical screening and RNAi to identify host genes involved in M. tuberculosis and Bacillus anthracis infection. We are also studying the pathogen-host interaction where heterogeneity exists in the interactions between immune cells to pathogens on the single cell level. At the other extreme, we are exploring host genes required at the whole organism level that are important for infection using zebrafish (Danio rerio) embryos as a model host and chemical screening for compounds that rescue an animal from lethal infection.

2) Identifying essential functions and small molecule tools with which to probe their in vivo biology. Bacterial functions required for survival during infection can be very different than those required for survival within a test-tube. Further, the complex physiology adopted by bacteria under certain conditions including within the host can allow them to persist in the face of host immunity and antibiotic treatment. We are developing novel approaches to identify small molecules that target essential functions of M. tuberculosis and P. aeruginosa under different growth and survival conditions that incorporate novel chemical biological and genomic technologies. We have developed custom barcoded strain libraries for comprehensive genetic and chemical screening to identify essential functions under these different conditions. Further, we have developed small molecules identified to target bacteria under these different conditions and are using them to probe in vivo biology in whole organism models of infection.

3) Identifying transcriptional responses to antibiotic stress and developing new tools to both rapidly diagnose a pathogen and determine antibiotic susceptibility based on these responses. Current diagnostic tests for infectious disease rely on culture-based methods, immunoasssays, and molecular techniques. These techniques are too slow, particularly in the case of M. tuberculosis where culture-based methods used to diagnose and determine antibiotic susceptibilities require weeks to months. In fact, bacteria alter their transcription in response to antibiotic exposure within minutes in efforts to evade the toxic effects of the antibiotic. By studying these rapid transcriptional responses, we are developing a sensitive and rapid approach to both identify a pathogen and determine its antibiotic sensitivity.