ABSTRACT
Globally, approximately 2.8M people are infected with bacteria that are deemed resistant to clinically relevant
antibiotics an annual basis. Of these infections, 700,000 individuals will die with the United Sates accounting
for 5% (35,000) of these deaths. In the 2019 report, The World Health Organization states that if no action is
take, drug resistant diseases could cause 10 million deaths each year by 2050 and that by 2030, antibiotic
resistance could force up to 24 million people into extreme poverty. There is an urgent need for the
development of novel antibiotics to combat the drastic rise in the number of antibiotic resistant bacteria. In
particular, there are few molecules with broad spectrum activity against multidrug resistant pathogens. One of
the bottlenecks that is impeding the further development of novel antibiotics is the lack of identified gene
clusters producing them, which could lead to optimized heterologous production. The overarching goal of this
proposal is to identify and characterize the biosynthetic gene clusters and products from these anabolic
pathways that responsible for the production of antibiotics, which in crude extracts were shown to be active in
killing multi-drug resistant pathogens. The proposed research is significant since antibiotic resistance to
clinically relevant antibiotics is increasing across different types of pathogenic bacteria, and particularly,
broad-spectrum options to treat multidrug resistant strains are currently limited. The PI and his students
recently isolated and identified several strains of rhizospheric bacteria, which possess broad-spectrum
antibiotic activity. These initial experiments were performed using a culture-based approach, which screened
for strains competing against each other on solid media, leveraging the chemical ecological concept of
competition between bacterial species from the same environmental sample. One of the strains,
Exiguobacterium sp RIT 594 produces a cocktail of antibiotic compounds (in response to another isolate
Acinetobacter sp RIT 592). The antibiotic activity produce a >6 log reductions in three relevant resistant
clinical isolates: (1) the epidemic strain MRSA USA300 (FPR3757), (2) NDM-producing E. coli (MCR1_NJ)
and (3) VIM-producing P. aeruginosa (AR-0266) in 24 h time kill assays. Importantly, the preliminary data
show that the emergence of in vitro resistance was not detected in the time kill assays. Subsequent analysis of
crude extracts using LC-MS provided evidence and identified more than 200 novel compounds that are
unknown in the mass spectrometry databases. Whole genome sequencing and annotation, revealed 8 putative
gene clusters that contained a large number of uncharacterized proteins that are predicated to produce a
variety of secondary metabolites that are putatively endowed with antibiotic properties.
