A Year in the Lab: Literature Week

I have reached a point in my identification where I really needed to sit down and work through all the data I have gathered over the last few weeks. So I decided to take a week out of the lab to focus on processing and interpreting my data, as well as working on the first part of my thesis: Literature Review and my GS3 form.

Monday 11th November

I started my week by visiting the lab to see whether my Kovac’s reagent for indole testing had arrived. Unfortunately it hadn’t, so I decided to have a run over the Spiral Plater to ensure that all the correct parts and accessories were present. I found a few pieces missing, however after speaking to my supervisor I discovered he had taken them home to check them over. As they were all accounted for, I left the lab and decided to spend the rest of the day focusing on inputting all my collected data into a database spreadsheet.

IMG_3540Spiral Plater

I decided to start with four main headings: Sample, Agar Growth, Biochemical Testing and ESBL Testing. I chose these headings as they incorporated the main areas in which I have conducted research and collected results.

Sample

This section included information relating specifically to the original samples stored in glycerol that make up the bacterial collection. In the following categories: Number, Suspected organism, Colour and Ability to pipette.

67, 68, 72Samples stored in glycerol

Number: The number is the numerical code that I found on each of the individual samples in the bacterial collection.

Suspected organism: When I first inputted my collected data I didn’t complete the suspected organism section, however later in the week I used the combined data I collected to determine this.   

Colour: When processing all the samples within the collection I made sure to take note and photograph each sample. I used this data to describe the colour of each sample.

Ability to pipette: One main thing I noticed when processing the samples, by spread and then streak plate, was that some samples were more difficult to pipette/work with. Again, I made sure to take note when this occurred, as I was unsure as to whether it may effect the final results of the procedure/tests for the sample.

Agar Growth

This section included information regarding whether each sample grew on MacConkey agar and the colony characteristics of those which did – originally by spread plating and then streak plating. I also heat fixed slides of samples and Gram stained them to observe cell morphology.

IMG_3653Two spread plated samples and two streak plated samples on MacConkey agar

MacConkey spread plate: While processing the samples I made detailed notes and took photographs of all plates. Whether the sample grew on MacConkey and if so whether the sample was lactose positive or negative.

Colony characteristics: If the sample did grow on MacConkey, I made notes of the individual colony characteristics. Most specifically – Shape, size, colour and appearance.

MacConkey streak plate: I also decided to streak plate some of the samples which produced excessive growth when spread plated. Again, I recorded details of the growth and colony characteristics of each sample.

Gram stain: As I expected all samples I processed were Gram negative, demonstrating a pink colour under light microscopy.

Cell morphology: Once Gram stained I observed the samples under light microscopy in order to identify the morphology of cells. Most samples demonstrated Rod shaped cells, in either pairs or clumps. Some appeared shorter and wider, while others longer and thinner.  

66 - x1000 mag (2)Gram stained sample – demonstrating Gram negative rod shaped cells

Biochemical Testing

As I described in my last post I decided to conduct three biochemical tests to help differentiate between bacteria. These included: Indole, Oxidase and Motility. So far I have only completed oxidase and motility tests, so I filled in these sections and will be completing the rest when I can conduct indole testing on my samples.

Indole: As I haven’t performed this test yet I left this section blank. When completed, samples will either be positive or negative.

Oxidase: When conducting oxidase testing I carefully recorded the result for each sample. I used this information to fill in whether a sample was positive or negative.

Motility: While performing motility testing I carefully noted down the result for each sample. I then used this information to fill in whether a sample was motile or non motile.

ESBL Testing

Although I didn’t conduct ESBL testing for this collection of samples, I plan to do so for future sample collections. I have been given this information from the previous MSc student who worked on this collection. This section includes information regarding the presence of the three most common beta lactamase genotypes: CTX, TEM and SHV. Extended spectrum beta lactasmases (ESBLs) are hydrolytic enzymes produced by a variety of Gram negative bacteria (Like my collection) that make them resistant to cephalosporins, penicillins and monobactams.

Tuesday 12th November – Friday 15th November 

I continued processing information and inputting results into my database spreadsheet for the rest of the week. Throughout this process I also worked through the many photographs I have taken of samples during the stages of identification. I combined images to allow comparison and easier interpretation. As well as this, I also spend time working on both my thesis literature review and GS3.

Bacterial database spreadsheetBacterial database spreadsheet

78 - Escherichia coliComparison of sample 78 spread plated and then streak plated

Literature Review

As I described in my 3rd year Biomedical Science Dissertation Project blog, a literature review focuses on the literature that enables the reader to get a better understanding of the basis and main aims of the research. While writing this section I researched and read many journal papers that contained information and results fundamental in the focus of my work.

GS3

In my ‘Forms, forms, forms’ post I mentioned the purpose and sections of the GS3 form – as it is nearly December I am finalising the details of this form. The process of researching and writing my literature review was very useful in the important/key references section of my GS3 form. I would definitely recommend doing these two stages together, as it allows you to really use up to date and key references in both your thesis and confirmation of study form (GS3).

GS3

Next week…

I am really glad I decided to take this week out to focus on my literature work. It has really helped to have all my data in one place, as it has allowed me to easily and quickly compare and interpret my results so far. Next week I hope to conduct indole testing on the samples as well as any tests that haven’t been completed for certain samples – due to various reasons. All samples within the collection now have a suspected organism in the ‘suspected organism’ section, however this won’t be fully confirmed until my final testing has occurred. I also hope to start setting up, cleaning and running samples with the Spiral Plater.

A Year in the Life…

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