Isolation and Characterization of Antibiotic Producing Soil Bacilli from Southern CA

The lack of new antibiotics together with the rise of multidrug resistant “superbugs” has led to an increase in mortality rate related to infections. The ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) are responsible for a substantial percentage of nosocomial infections in the modern hospital, and represent the vast majority of isolates whose resistance to antimicrobial agents pose serious therapeutic dilemmas for physicians. Soil bacteria, particularly from the genera Bacillus and Pseudomonas, produce a large variety of secondary metabolites with antibiotic activity that not only protect them from other microbes, but also play an important part in quorum sensing, biofilm formation, interactions with plants, and sporulation. The Small World Initiative (SWI), spearheaded by Yale University, integrates the search for novel antibiotics into the undergraduate science curriculum. The goal of the SWI is to isolate undiscovered antibiotic-producing bacteria from soil samples through undergraduate research, effectively “crowdsourcing” antibiotic discovery. As part of National University’s Introductory Microbiology course, we plated soil samples from diverse locations in Orange and San Diego counties. Colonies were tested for antibiotic production using spread/patch technique against safe surrogates of the  “ESKAPE” organisms. Cultures exhibiting antibiotic production were further characterized using a combination of biochemical and genetic techniques. Most students in the class obtained at least one antibiotic producer. We report three Bacillus species obtained with a diverse range of activity against ESKAPE organisms. Biochemical, morphological, and genetic tests suggest the obtained species are probably related to B. mojavensis or tequilensis, although more sensitive methods such as fatty acid analysis or whole genome sequencing are needed for a definitive identification. Preliminary chemical fractionation results indicate that multiple active compounds are present in both organic and aqueous fractions. In summary, the approach described constitutes an effective and straightforward way to obtain a pipeline of possible antibiotics while providing a classroom research experience.