A Wild Pseudomonas has appeared: An Exercise in Bacterial Isolation and Identification

The aim of our research was to isolate and identify wild type Pseudomonas putida from soil in various cities of San Gabriel Valley. P. putida iscapable of biomineralization. Biomineralization can potentially be used as a method of phosphorus recovery by using bacteria to produce phosphate rich struvite. In isolating bacteria for further observation, fluorescence was used as a primary determinant in identifying possible Pseudomonas strains as fluorescence is a common trait shared among varying Pseudomonas species; P. putida, P. aeruginosa, P. fluorescens, P. cichorii, P. chlororaphis, P. syringae,and P. aureofaciens. King’s B agar was used to promote the production of pyoverdine in these strains (allowing for direct identification based on a green fluorescence under UV light) as this medium has specific ingredients that enhance pigment production.These fluorescent bacteria were then further isolated from each other and identified using biochemical methods including catalase, oxidase, nitrate reduction, and gelatin hydrolysis tests to differentiate P. putida from the six other fluorescent Pseudomonas species. Of the 21 total samples isolated based on fluorescence, 5 of the samples were determined to be potential P. putida. While the biochemical assays were conducted, the isolated samples were placed in refrigeration for 3 weeks. After the biochemical tests were completed and 3 weeks had passed, visible crystals had formed in the potential P. putida. Albert’s metachromatic staining was performed to determine the presence of polyphosphate granules. Ultimately, each potential P. putida that produced crystals also showed polyphosphate granules when Albert’s stained, which further connects crystal formation with prior polyphosphate formation