{"id":707,"date":"2020-03-11T18:15:39","date_gmt":"2020-03-11T18:15:39","guid":{"rendered":"https:\/\/blogs.ncl.ac.uk\/bns\/?p=707"},"modified":"2022-03-21T20:40:14","modified_gmt":"2022-03-21T20:40:14","slug":"my-summer-with-cryptosporidium","status":"publish","type":"post","link":"https:\/\/blogs.ncl.ac.uk\/bns\/my-summer-with-cryptosporidium\/","title":{"rendered":"My Summer Travels with Cryptosporidium"},"content":{"rendered":"\n

By Rosie Gathercole<\/h3>\n\n\n\n

Working with poo turned out to be exactly the summer experience I wanted! <\/p>\n\n\n\n

I worked at the national Cryptosporidium <\/em>Reference Unit (CRU) at Public Health Wales in Swansea with Professor Rachel Chalmers and her team. I received a Scholarship from the Society for Applied Microbiology (SfAM) for this placement, writing the application together with Rachel.<\/p>\n\n\n\n

\"Rosie
Me working on my summer placement<\/figcaption><\/figure><\/div>\n\n\n\n

Cryptosporidium <\/em>is a parasite that causes diarrhoea, is found globally\nand is typically passed from animals, other people, food and fresh water\nsources. It is currently a human health issue due to the significant effect it\nhas in developing countries and the lack of specific treatments to fight the\nparasite. Quite often how well you recover from the illness depends on how\nhealthy you were to begin with!<\/p>\n\n\n\n\n\n\n\n

What did I actually do in<\/strong> Swansea?<\/strong><\/h3>\n\n\n\n

After reviewing the literature, we quickly realised that it would\nbe very useful if we could analyse the protein content of Cryptosporidium <\/em>because\na clear understanding of proteomics can help the current genomics work being\ndone in the area. To do this the parasites would first have to be isolated from\nhuman faecal samples at the CRU. <\/p>\n\n\n\n

\u201cThe CRU gets sent lots of samples, yes that means lots of parasite filled poo!\u201d <\/strong><\/p>\n\n\n\n

My project was to optimise a way to isolate parasite \u201ceggs\u201d from these\npoo samples ready for protein analysis. What I ended up doing was performing\n\u2018salt flotations\u2019 initially to separate the parasites from the faeces. This\ninvolved placing the samples on salt-water and spinning in a centrifuge. Next, I\nused potassium bromide solutions to separate contents specifically based on\ntheir density. However, the more you try and \u2018clean-up\u2019 a sample, the more\nparasites you lose from it so it became a bit of a balancing act which involved\nfiddling with centrifuge speeds, volumes of samples, and reagents for a week or\ntwo.<\/p>\n\n\n\n

\"Cryptosporidium
My samples on potassium bromide gradients, ready and waiting to be centrifuged<\/em><\/figcaption><\/figure><\/div>\n\n\n\n

This photo shows the samples (still a lovely brown colour) on top\nof the potassium bromide solutions. After spinning you can now see a white band\ncontaining the parasite eggs. This band was then extracted and washed.<\/p>\n\n\n\n

\"A
A successful sample straight out of the centrifuge. The white band contains the parasites.<\/em><\/figcaption><\/figure><\/div>\n\n\n\n

It worked!<\/strong><\/h3>\n\n\n\n

In order to count the number of eggs that I could purify using\nthis method, I pipetted the extracted samples onto a tiny grid on a microscope\nslide. I was hoping to be able to purify at least 1 million eggs in order to\nhave sufficient material for the next step so I was ecstatic to come across a\nsample with over 10 million!<\/p>\n\n\n\n

\"A
A light microscope image of extracted Cryptosporidium \u201deggs\u201d. Can you spot the empty shell? <\/em><\/figcaption><\/figure><\/div>\n\n\n\n

Each \u201cegg\u201d that can be seen contains four Cryptosporidium <\/em>parasites,\nso my sample, actually contained more than 40 million parasites. All purified\nin around 1ml of clear liquid.<\/p>\n\n\n\n

After collecting as many good samples as I could, I presented my protocol and results to the CRU team<\/a> to show them that I had been successful since letting me loose in their lab.<\/p>\n\n\n\n

Analysing my purified parasites <\/strong><\/h3>\n\n\n\n

Next, I suspended the parasites in ethanol to kill any contaminants,\npackaged them in dry ice, and drove to Liverpool University Veterinary School where\nI was lucky enough to work with Erika Transfield, a MALDI application\nspecialist for the Bruker Corporation. <\/p>\n\n\n\n

\"The
The mass spectrometry machine in Liverpool, manufactured by Bruker. <\/em><\/figcaption><\/figure><\/div>\n\n\n\n

My aim at Liverpool was to analyse my purified Cryptosporidium <\/em>samples using their latest MALDI-TOF<\/a>\nmass spectrometer. Mass spectrometry machines are able to tell you what the\nmass for every bit of protein in your sample and from this you can determine\nwhich proteins are present. This had never been performed successfully with Cryptosporidium\n<\/em>before, but we were very hopeful that the developed protocol would be\nsuccessful.<\/p>\n\n\n\n

First, I added a bunch of chemicals to my samples to \u201cfix\u201d the\nproteins and then I pipetted them onto a metal plate that would go into the\nmachine.<\/p>\n\n\n\n

\"Rosie
Me pipetting my samples onto the plate that then goes into the mass spectrometer machine.<\/em><\/figcaption><\/figure><\/div>\n\n\n\n

Success!<\/strong><\/h3>\n\n\n\n

What you can see on the computer screen behind me was the first\nsuccessful protein peaks (spectrum) that the computer spat out. After a few\nhours we ended up with two successful spectra from the six samples that I had brought\nwith me from Swansea. Despite being back at university for my final year, I am\nstill in contact with the team as we analyse the data to identify the specific\nprotein fragments the mass spectrometer identified.<\/p>\n\n\n\n

\"Rosie
Me at the computer with my first spectra. The results were being generated live onto the screen.<\/em><\/figcaption><\/figure>\n\n\n\n

On to Aberystwyth University <\/strong><\/p>\n\n\n\n

Straight after generating this exciting data, I then was lucky enough to be whisked away to Aberystwyth University where I spent two more weeks in the bioinformatics office with a PhD student who showed me how to work with massive data sets. We used operating systems such as Linux to analyse the complex data generated from these parasites. We used scripts to see if any of my samples were unique e.g. If one sample contains proteins that none of the others did, that could be a way to identify that subtype of Cryptosporidium<\/em>?<\/p>\n\n\n\n

\u2026.and finally<\/strong><\/h3>\n\n\n\n

” I would definitely recommend doing a placement over the summer”<\/strong><\/p>\n\n\n\n

This experience as a whole was able to provide me with a seemingly endless list of transferable skills which I am already finding to have transformed the way I approach my 3rd<\/sup> year studies. Also being able to work so freely within a research-based work environment has fuelled my interest in lab-based science and is certainly influencing my post-graduation plans. Being able to work side-by-side with a talented bioinformatician in particular has kick-started my passion for the field and I am still continuing to play with the software I was introduced to just a few months ago.<\/p>\n\n\n\n

I would definitely recommend doing a placement over the\nsummer and I am incredibly grateful to Professor\nRachel Chalmers<\/a> for taking the time to organise and oversee my\nproject, her team for helping me in the lab and SfAM<\/a>\nfor the funding.<\/p>\n\n\n\n

Rosie did\nthis fantastic work in the summer between her second and final year of BSc Biomedical Science<\/a>\nin the School of Biomedical, Nutritional\nand Sport Sciences.<\/a><\/p>\n\n\n\n

Read about\nsummer placements our other students have done here<\/a>.<\/strong><\/p>\n","protected":false},"excerpt":{"rendered":"

By Rosie Gathercole Working with poo turned out to be exactly the summer experience I wanted! I worked at the national Cryptosporidium Reference Unit (CRU) at Public Health Wales in Swansea with Professor Rachel Chalmers and her team. I received a Scholarship from the Society for Applied Microbiology (SfAM) for this placement, writing the application … <\/p>\n

Continue reading “My Summer Travels with Cryptosporidium”<\/span><\/a><\/p>\n","protected":false},"author":2915,"featured_media":714,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[168,82,86],"tags":[113,63,17,18,73,58,126,93],"class_list":["post-707","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-all-posts","category-placements","category-summer-placements","tag-biomedical-sciences","tag-employability","tag-research","tag-student-projects","tag-student-voice","tag-studentsuccess","tag-summer-placement","tag-work-experience"],"_links":{"self":[{"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/posts\/707","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/users\/2915"}],"replies":[{"embeddable":true,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/comments?post=707"}],"version-history":[{"count":12,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/posts\/707\/revisions"}],"predecessor-version":[{"id":1431,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/posts\/707\/revisions\/1431"}],"wp:featuredmedia":[{"embeddable":true,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/media\/714"}],"wp:attachment":[{"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/media?parent=707"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/categories?post=707"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/blogs.ncl.ac.uk\/bns\/wp-json\/wp\/v2\/tags?post=707"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}