BioTech Lab Enrty 3

 Week Three

Introduction

   This week we innoculated several medias and performed biochemical characterization tests on Deinococcus xinjiangensis. 

Methods

    A gram stain is done on the D. xinjiangensis. They test gram -. The plates labeled last week are innoculated with their respective bacteria species. The tubes containing SIM media are innoculated by opening the tube and passing it over a flame. The smaller tube containing the bacteria is then swabbed. The needle is then dipped into the larger tube, which is passed over the flame again and closed. The methods of using UniProt and NCBI to detect the presence and absence of genes involved in chemotaxis/mechanotaxis is then scrutinized. It is discovered that the method we had been using was unreliable. Following this, two flasks of TGY are innoculated with D. xinjiangensis. The following day, we examine the plates for growth and inoculate the citrate plates with D. xinjiangensis. A rough draft of the abstract is started and edited. The importance of studying Deinococcus as a genus is then discussed, as is the information which goes into an abstract. The following day, biochemical characterization tests are performed on Deinococcus xinjiangensis, which I am not present for. However, according to the protocols used, this is how the tests proceeded. 1 drop of 1% Kovac's reagent was placed on the plates for the oxidase test. Urease plates are examined for the coloration after being streaked the day prior. The SIM tubes are examined for the presence of black coloration. Kovac's reagent is added to the solutions, and the tubes are innoculated with their respective bacteria using an innoculation needle. 2-3 drops of 10% iodine are added to the edges of the starch plates and they are carefully observed. The following day. The following day, further gram stains are performed (which give negative results) and we continue to work on the abstract. 

Results

    The oxidase and indole tests were inconclusive. Deinococcus xinjiangensis tested catalase +, urease -, motility -, sulfur-reduction -, and amylase  +. 

Discussion

    Oxidase and SIM tests must be re-done in order to get results. In the oxidase test, the reagent turned everything blue. However, I do not think that this is the result of contamination or an issue with the reagent. Kovacs reagent will oxidize naturally, so it is the speed at which the plate turns blue which determines whether or not a given species is positive or negative for the result, and not the presence or absence of the color blue. It is worrying that all of our gram stains have been negative, given that the bacteria are supposed to test positive. However, I am septical that this is the result of contamination, as that usually means there is a mix of both gram positive and gram negative bacteria. Due to this I am more inclined to doubt the validitiy of the initial claim, but I remain open to either possibility. The phylogeny of D. xinjiangensis deeply interests me. According to Peng et al. 2009, D. xinjiangensis is sister to D. hopiensis. However, more updated phylogenies dispute this. I am incredibly interested in what the true phylogeny for Deinococcus is, if that can even be determined. I am very interested in whether or not the discrepencies in phylogenies are the result of horizontal-gene transfer (HGT). How can we account for HGT when reconstructing phylogenies? Is it even possible? 


References

    “1.23: Sim Deep Tests.” Biology LibreTexts, Libretexts, 3 Mar. 2023, bio.libretexts.org/Bookshelves/Microbiology/Microbiology_Laboratory_Manual_(Hartline)/01%3A_Labs/1.23%3A_SIM_Deep_Tests.

Aryal, Sagar, and Wilhelmina.Debbie. “Urease Test- Principle, Media, Procedure and Result.” Microbiology Info.Com, 10 Aug. 2022, microbiologyinfo.com/urease-test-principle-media-procedure-and-result/.

“Oxidase Test Protocol.” American Society for Microbiology, asm.org/getattachment/00ce8639-8e76-4acb-8591-0f7b22a347c6/oxidase-test-protocol-3229.pdf. Accessed 31 Mar. 2024.

Peng, F., et al. “Deinococcus Xinjiangensis sp. nov., isolated from desert soil.” INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, vol. 59, no. 4, 1 Apr. 2009, pp. 709–713, https://doi.org/10.1099/ijs.0.004564-0.

Rijal, Nisha. “Starch Hydrolysis Test: Principle, Procedure, Results • Microbe Online.” Microbe Online, 29 Dec. 2023, microbeonline.com/starch-hydrolysis-test/. 

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