Seek and you shall NOT find.

Bacteriophages are viruses that infect bacteria. They are highly abundant and were discovered around 100 years ago but there unrealised potential is only being discovered now. In the future, bacteriophages may have the possibility to advance health care and medicine, especially when it comes to the antibiotic resistance crisis. This is why they are of immense interest to the scientific community as they could revolutionise modern medicine. Antibiotic resistance is becoming a huge problem in the modern world as illnesses and infections cannot always be cured by antibiotics now. Phage therapy is an alternative to using antibiotics and could be used to save millions of lives. In the United States alone, more than 23000 people die every year due to antibiotic resistance.[1]  If you want to find out more about phage therapy, you can do so here.[2]

Bacteriophages are the most abundant organism [3] on the planet and so there are potentially millions of bacteriophages in a compost bin and yet they all evade me. Taking environmental samples to find bacteriophages is the first step to phage hunt. This shouldn’t have been that hard considering that there are trillions of bacteriophages in a garden let alone around the world. I thought that this would’ve been the easiest step but I was so wrong. It was so much harder than I thought as I took 27 soil samples from locations around the North Shore and didn’t find a single bacteriophage. I collected the samples in batches of 10, each round taking about an hour to finish. I looked in compost bins and gardens and worm farms and streams and paddocks hoping to find these elusive phages. I started to think that we had been told wrong as these things that we were searching for were mean to be in such high abundance that it should’ve been easy to find them. By the end of it I was sick of collecting samples in plastic tubes only to find out after the first round of processing and plating that I hadn’t found anything. Links to some of the protocols we followed can be found here and here. The second link is similar, but not identical, to a protocol that we followed.[4, 5]

I was considered the unluckiest phage hunter in our lab by our supervisors, sampling for a solid three weeks with no phages found after first round processing. Some of my co phage hunters were lucky enough to find phages in their first round of samples and yet they remained in hiding for me. The graphs show the number of environmental samples taken before people found or adopted a phage. The positive graph shows those who found phages and the negative, obviously, shows those who were unsuccessful at finding phages. There didn’t really seem to be any relationship between finding a phage and the number of samples taken. It seemed to come down to a bit of luck and looking in the right places.

Negative phagepositive phage

The phages we were looking for were ones that infected the bacteria Mycobacterium Smegmatis. If the phages infect Smegmatis then they also have the potential to infect Mycobacterium Tuberculosis. M.T is a close relative to M.Smeg and so there is the hope that our experiments and research could lead to finding a phage that could be used in fighting Tuberculosis. It could potentially save the lives of millions of people and help improve modern medicine and move past the overuse of antibiotics. With antibiotic resistance becoming an ever increasing problem in our world, this kind of research can possibly contribute to the ongoing scientific research of bacteriophages. The diversity and abundance of bacteriophages is immense and so there is still so much to learn about them and the possibilities they can bring.

Mycobacterium Smegmatis is said to be found in places with lots of nutrients / water sources which is why I looked in places that had an abundance of one and or the other. I collected all of my samples from places like this and hence my confusion and frustration when I didn’t find any!

Due to my inability to find phages, I had come to the end of a month of searching with no success. I was lucky enough to adopt a phage from my friend. She had managed to find three in one sample. Things were going great and by week six I was up to making a high titre lysate of my tiny little phage. A link to what protocol we followed can be found here. [6]  I was beginning to think that I might have gotten a handle on phage lab but as always I managed to lose the phage I had adopted. I had nicknamed my phage “Moppet” as it formed small plaques. My phage was not surviving in phage buffer so I had to go and adopt another phage from one of my other amazing co phage hunters. This meant that I had to redo the webbed plating and making a high titre lysate processes. It may have taken a long time to get to this stage but I got there in the end and had found a bacteriophage.

Hunting for bacteriophages is a tricky business but once you find a phage that you have spent tedious amounts of time finding and purifying and amplifying it all becomes worth it. While my hunt for phages didn’t result in finding my own phages, I was still able to come out with a phage that will hopefully see me through to the end and you can’t say I didn’t try.

Stay tuned for the next instalment.

References

  1. Antibiotic Resistance Threats in the United States, 2013.
  2. What is Phage Thearpy.
  3. Martha RJ Clokie, A.D.M., Andrey V Letarov, and Shaun Heaphy, Phages in nature. Bacteriophage, 2011 Jan-Feb.
  4. Biosciences, B., Plaque Assay
  5. Trevor Cross, C.S., Dylan Chudoff, LIbby Graves, Haley Broomell, Katrina Terry, Jennifer Farina, Alexandra Correa, David Shade, and David Dunbar An Optimized Enrichment Technique for the Isolation of Arthrobacter Bacteriophage Species from Soil Sample Isolates.
  6. Phagesdb, Manufacturing a high titre lysate.

 

Advertisements
This entry was posted in Uncategorized. Bookmark the permalink.

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out / Change )

Twitter picture

You are commenting using your Twitter account. Log Out / Change )

Facebook photo

You are commenting using your Facebook account. Log Out / Change )

Google+ photo

You are commenting using your Google+ account. Log Out / Change )

Connecting to %s