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

Abstract

<span id="docs-internal-guid-8e962b78-7fff-c5ee-bc79-1ba2a6c149e3"><span>The aim of our research was to isolate and identify wild type </span><span>Pseudomonas putida </span><span>from soil in various cities of San Gabriel Valley. </span><span>P. putida </span><span>is</span><span>capable 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 </span><span>Pseudomonas </span><span>strains as fluorescence is a common trait shared among varying </span><span>Pseudomonas </span><span>species; </span><span>P. putida, P. aeruginosa</span><span>,</span><span> P. fluorescens</span><span>,</span><span> P. cichorii</span><span>,</span><span> P. chlororaphis</span><span>,</span><span> P. syringae</span><span>,</span><span>and </span><span>P. aureofaciens</span><span>. 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.</span><span>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 </span><span>P. putida </span><span>from the six other fluorescent </span><span>Pseudomonas </span><span>species. Of the 21 total samples isolated based on fluorescence, 5 of the samples were determined to be potential </span><span>P. putida. </span><span>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 </span><span>P. putida</span><span>. Albert’s metachromatic staining was performed to determine the presence of polyphosphate granules. Ultimately, each potential </span><span>P. putida</span><span> that produced crystals also showed polyphosphate granules when Albert’s stained, which further connects crystal formation with prior polyphosphate formation.</span></span>