The US Centers for Disease Control and Prevention (CDC) have identified multi-drug resistant (MDR) pathogens (e.g. the six ESKAPE pathogens) as a significant and growing threat to human health due to their recalcitrant and increasing antibiotic resistance. In particular, methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of morbidity and mortality in hospital-acquired infections. Over the last three decades, the search for new antibiotics from natural or synthetic small molecule libraries has met very limited success. Antimicrobial peptides have emerged as a promising alternative or complement to chemical compounds in treating bacterial infections. Based on our experience and limited, but promising, evidence in the literature, we posit that organisms from diverse environmental habitats produce peptides and proteins with potent anti- microbial/anti-biofilm activity. Our central hypothesis is that peptides with antimicrobial properties may be efficiently isolated from small- insert DNA libraries created from soil, human and/or animal microbial communities using a novel ultra-high- throughput (uHTS) nano-scale cellular microarray. Specifically, we will (i) integrate functional metagenomics and nanoculture to develop a `Library on a chip' consisting of a million unique clones on a single slide; (ii) interface the `Library on a chip' with a `Pathogen on a chip' nanoculture assay to prospect for peptides with antimicrobial activity including bactericidal and/or anti-biofilm effects against pathogenic S. aureus; (iii) annotate the sequences, and validate their proteolytic stability, toxicity, and activity against MRSA clinical isolates. Upon successful completion of this work, we will have developed an uHTS platform for creating metagenomic `libraries on a chip' that interface seamlessly and modularly with `pathogen on a chip' bactericidal or anti-biofilm assays. We will have tested the hypothesis that microbes naturally produce peptides that interact pathogens to influence cell growth, viability and/or biofilm production; thus delivering novel compounds with therapeutic potential, and as chemical probes to understand virulence and survival. In a larger context, our screening platform may be easily tailored not only to target virulence mechanisms associated with other bacterial infections but also discover peptides with heretofore unknown functions for a number of biomedical applications.
Bacterial infections represent a significant healthcare burden across the globe, driven primarily by increasing resistance to traditional antibiotics and reduced discovery of new antibiotics, and therefore requiring non-traditional approaches to combat these serious threats. To this end, we will identify peptide molecules against growth, viability and biofilms of methicillin-resistant Staphylococcus aureus (MRSA) by screening genes from uncultured soil, animal, or human microbial communities using a novel ultra-high-throughput nanoculture platform. This approach is fundamentally different from traditional discovery programs both in the content of the library being screened (metagenomes are evolutionarily enriched for peptides with large structural and functional diversity) and in the screening methodology (iterative screening at nano-liter volume for rapid selection) which combine to maximize the potential for discovering functional peptides not only as antibiotics but also for a wide range of biomedical applications.
