The contents of this page are the words and images of Natasha Boyd, a Nuffield Foundation student from Winstanley College, Wigan, undertaking a 1-month research placement in my lab in August (2014). Natasha has come to MMU with no experience of microbiology. Below is a diary of her time here, written in her own words. Images are the property of Natasha Boyd.
Week 1 Diary
Day 1 – 4/8/14
I arrived at MMU at 9:30 with a mixture of excitement, nerves and anticipation. I have been looking forward to this placement for months and yet I am a little daunted at the prospect of working alongside accomplished scientists. When I was waiting in the lobby I met with some lovely girls who were starting their projects today also and the fact that they were also a tad apprehensive calmed me a little.
I met with Conway – the Nuffield coordinator for MMU. His cheery disposition soon put me at ease and he explained how the project worked and what I could expect from my placement. We then went on a tour of the campus with Manny, a first year environmental science student at MMU.
I then had an introduction to the microbiology department followed with Ann, Technical Manager. We discussed the health and safety protocols (e.g. fire drills) and how to work in a safe and environmentally friendly way in a lab. The tour of the microbiology labs was a great chance to view all the equipment that microbiologists use on a daily basis – they are nothing like the labs at my college. Having never studied microbiology before I am embarking on a very steep learning curve!
I then talked to Laura about my project, it will be based on antibiotic resistance of microbes in the Irwell River and so has a slight connection to the work Laura is doing with Ian, a researcher in Geography and Geography at the university. We also discussed the fieldwork that we would be doing the next day -off to wade in a river tomorrow.
Day 2 – 5/8/14
This morning I donned wellies and went on a field trip to the Irwell River with Laura and Ian. We took both samples and measurements at the field site. Samples of the water and sediment were taken from both the right and left hand side of the bar. Measurements were taken of the amount of dissolved oxygen, pH, the temperature and the rate of flow (m/s)
- The importance of aseptic techniques (e.g. working around a Bunsen burner and sterilising all apparatus and surfaces) in microbiology in order to avoid contamination
- The different types of isolation techniques and what they are used for
- Spread plate
- Streak plate
- Pour plates (which I will not be using)
- The technique of serial dilution whereby one takes 1ml of the original sample and add it to 9ml of sterile water to create the 10-1 This process is then repeated to create a 10-2 solution and so on up to 10-6. This is to prevent the petri dish from being too over crowded to distinguish between different colony forming units (cfu). The ideal is to have 30-300 cfu on each dish.
- How to prepare a spread plate of the water samples we took (including how to successfully use a pipette).
- I then prepared some media (with the help and direction of Laura)
That evening Laura asked me to think about what factors affect microbe communities in a river. (below is what I came up with)
Day 3 – 6/8/14
I prepared some agar, but this time on my own! Laura then came up to the labs and showed me how to safely use the autoclave (which I learnt is essentially a fancy name for a big, hot pressure cooker). We then autoclaved the agar and some water (in order to sterilise them) at 15 pounds per square inch and 121°C for 15 mins.
Prepared my universals with 9ml in each in order to carry out serial dilution on my samples tomorrow.
Day 4 – 7/8/14
Today I had a very busy morning. I completed serial dilutions for samples of water and sediment
I completed 3 spread plates for each dilution of each sample. It is important to do 3 for each so that an average can be taken and any anomalies spotted.
First thing this morning I briefly went into the lab to check on my spread plates and bag them up in order to ensure they would not contaminate the bench or other surfaces in the lab as the petri dishes are to be incubated at room temperature on the bench. There were some signs of growth in the 10-1 dilution petri dishes for all four samples the most prominent of which was the sediment samples.
I then did some research to find some relevant research articles using search engines such as google scholar
Week 2 Diary
Day 6 – 11/8/14
The first task of the day was counting the colony forming units that were now present on my spread plates after 96 hours. I marked these so that I could see which new colonies had formed by the next time I counted.
I then made up yet more agar and poured it into Petri dishes ready for my streak plates the next day. I had some downtime in the afternoon and so researched how antibiotics work and looked into the resistance test for antibiotics (I’ll be using a MASTRING-S™ which is like a ring with antibiotic tips)
Day 7 – 12/8/14
A new technique today – streak plating. I carried out this technique 40 times!! I randomly selected 10 colonies from each sample and, using an inoculating loop, aseptically made the plates.
In the afternoon I did some research into how antibiotic resistance can be transferred within microbial communities through transfection, transduction and conjugation.
Day 8 – 13/8/14
Today was mainly a day to take stock and evaluate where I was with my project. I checked and documented my streak plates (one of them shown below) and then came up with a day by day plan of the remainder of my project – a bit dull but necessary.
Day 9– 14/8/14
A day off to celebrate A-level results!!!!!!!!!!!!!!!!
Day 10– 15/8/14
MORE AGAR!! This time I needed 80 plates (2 for each isolate). I also prepared some nutrient broth. As agar can go dry (making it therefore trickier to grow cultures on), I stored the plates in the chiller over the weekend.
I then prepared my informal presentation for Monday that is to be shown to the other Nuffield students at MMU, the MMU Nuffield Coordinator Conway and David Ward – the head of the Greater Manchester STEM Centre. The presentation is only two slides long and is on my project so far.
Week 3 Diary
Lauras note: here’s an up to date pic of Natasha this Friday afternoon, taken sneakily while she was observing her culture collection. As ever under the watchful eye of our kind lead technician Anne. Natasha has had a very busy week and is now starting to see the fruits of her labours, with results coming in and new ideas being formed. Pity the placement is only 4 weeks!
On Monday Natasha joined the other Nuffield Foundation students at MMU for unofficial ‘mid-term’ presentations, discussing their projects with each other and their coordinator.
Week 3 has got to be the busiest week yet! This was the week that I actually did all my testing.
I inoculated all 40 samples by adding the culture to a universal filled with 9µl of nutrient broth. I then left them overnight in an incubator on a shake plate to allow aeration. I then tested for antibiotic resistance by adding a solution of the cultured cells to an agar plate with an antibiotic disk. I then left the plates to grow overnight. I identified where there was antibiotic resistance by observing growth around the antibiotic disk head.
I also used my inoculations to test whether the bacteria I cultured from my original samples are killed by exposure to UV light. I placed 1ml of my incubations, which were by now turbid, into eppendorfs, and centrifuged in order to separate the broth from the cells. I then extracted the nutrient broth leaving behind the cells which I then washed with sterile water. I added each sample to separate wells on a 92 well plate. I then left the well plate exposed to a UV light and then extracted 10 µl and placed droplets on an agar dish after 5 minutes, 20 minutes and 1 hour. Growth during incubation over the weekend would suggest that the bacteria were resistant to that particular length of exposure time. I’ll collect my results next week.
Week 4 Diary
Lauras note: Natasha spent her last week of her Nuffield placement analysing her results from week 3, performing additional experiments testing her isolates resistance to ethanol, and getting to grips with microscopy following the gram-staining of her antibiotic-resistant isolates. A busy week, not only finishing lab experiments but also preparing her Nuffield report and her application for the Gold Crest Award (which I’m sure she’ll have no problem achieving). With barely time to catch a breath before she heads back to college for her A-levels, understandably Natasha has decided not to update her diary this week and devote her time to her reports.