Biochemical Tests for Characterization of Deinococcus xinjiangenis

Introduction

Deinococcus xinjiangensis is a faintly pink, coccus shaped bacteria isolated from the Taklamakan desert located in Xinjiang, China. This species is understudied , with one paper based on the characterization of this species in 2009. This paper found that D,xinjiangensis was gram-positive, spherical, aerobic, non-motile, pink-pigmented and sensitive to desiccation and gamma radiation but highly UV resistant. We are interested in extending off of this paper by using more biochemical tests and UV resistance tests to further characterize D.xinjiangensis. We would like to do this because often, grand ideas are found in the most understudied of species. For instance, CRISPRS were randomly found in E.coli in 1987. 

Biochemical tests may require making a special media that promotes a change in growth after inoculation. Others require a reagent that act as a visual cue to test for products of a metabolic reaction.  Some require both, but all test for biochemical reactions in the cell. 

The tests we're focusing on using are the amylase, urease, citrate, oxidase, catalase, and SIM tests , as well as UV testing. We would also like to compare these tests on TGY vs R2A plates.l


Methods

The starch hydration test will require making a starch and TGY or R2A. When streaked and grown , bacteria will digest the starch using amylase. When iodine is introduced, a dark violet color produces except for around the smear where the amylase was used to digest the starch.

The Catalase test using the enzyme catalase to breakdown hydrogen peroxide. Hydrogen peroxide breaks down into H₂O and O₂ gas. The presence of bubbles or O₂ gas is a positive for the catalase test. 

The Urea tests for the enzyme urease which breaks down urea. This requires a special plate of water and urea. When urea is digested with the enzyme urea, the plate changes color from a peach tone into a bright pink due to pH indicator phenol pink. 

The SIM test tests for sulfur reduction, motility and indole. The SIM test is a 3 in 1, and requires SIM media. Slants are made and bacteria is inoculated with a needle. Motility is determined by how far from the inoculation point the bacteria grow. The indole test uses Kovacs reagent to determine if the cell converts tryptophan into indole using tryptophanase. The sulfur reduction tests if the bacteria is able to use sulfur as a main carbon source into hydrogen sulfide gas. The agar will turn black frim the hydrogen sulfide gas as a positive result.

The Oxidase tests for oxidation during the electron transport system. This will be done on R2A and TGY as it is dependent on a reagent. The reagent used is tetramethyl-p-phenylenediamine dihydrochloride which acts as an artificial electron acceptor. Once oxidized, it will turn blue within 2 minutes if positive. 

The citrate tests for if bacteria can use citrate as a carbon source of energy that will use the pH indicator bromothymol blue when the plate turns alkaline. 


Lastly, the UV test tests the cells resistance to varying UV exposure. However, this will be done in the next week. 

The species we will be using as our controls are Bacillus subtilis, Citrobacter freundii, Escherichia coli, and Staphylococcus epidermis.


Results


Catalase positive result characterized by production of O₂ gas


Urease negative result due to pH indicator phenol pink not turning bright pink
Negative Citrate result due to pH indicator not turning blue
Oxidase positive result characterized by blue kovacs reagent
Indole negative, motility negative, sulfure reduction negative result due to yellow topping and no black agar or fanning

Amylase positive result characterized by halo around inoculation streak


Gram-positive stain of D. xinjiangensis



Discussion 

From the results, we've found that D.xinjiangensis is gram positive, amylase, and oxidase positive. We also found that D.xinjiangensis is non-motile and citrate, indole, sulfur reduction and urease negative. The urease test will be reattempted as it contradicts the results from the original paper in 2009. Furthermore, uv resistance testing still needs to be done in order to gain more insight on this strain. 


Sources

Peng, F., Zhang, L., Luo, X., Dai, J., An, H., Tang, Y., & Fang, C. (2009). Deinococcus xinjiangensis sp. nov., isolated from desert soil. International journal of systematic and evolutionary microbiology, 59(Pt 4), 709–713. https://doi.org/10.1099/ijs.0.004564-0

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