The Beauty of Science

When I hear that “scientists are working on a new weight loss treatment”, or “scientists have found that that bacteriophages can help fight the war against antibiotic resistance”, I don’t really think twice about what goes into these research projects. It’s only once I started doing my own inquiry-based lab work that I started to understand some tunnels seemed like they had no light at the end, and how much blood, sweat, and tears accompany the satisfaction of progressing in science.

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I took part in the SEA PHAGES program, an international university course that allows you to discover viruses that kill bacteria in some dirt that you collect, purify them and name them, then rip the DNA out and send it through enzymes that chops it up into little pieces so you can examine your “phage”.

Sounds reasonably simple, right?

Well, I collected eight samples, and found no phage at all. Nudda. It demotivated me to keep going, but I would’ve failed the course if I didn’t collect more.

Wrong mind set, I know.

The next soil sample had amazing results: the phage had eaten all of the bacteria I had fed it! I was pretty stoked. But, after I started purifying and pulling the DNA out of my little phage (I called it “Mushball”, after my nickname), problem after problem occurred. When I added enzymes to cut the DNA, my DNA was gone.

That’s when the opportunity arose. I repeated the whole process, changing up little things in the hope that I got something. I ended up repeating it at least seven times, with the last time was the best result I could’ve got! I was ecstatic. So much frustration and perseverance had finally paid off, and the light at the end of the dark tunnel shone bright for me.

Science takes many turns that you would never have expected, and no matter how many repetitions you do, or how closely you follow the protocol to a “T”, you can never avoid a sigh of disappointment, a cry in the bathroom, or a broken piece of equipment you just threw at a wall. You begin to accept this, which makes it so much more exciting!

Who wants to discover something that does exactly what you expect it to?

“We learn wisdom from failure much more than from success; we often discover what will do, by finding out what will not do; and probably he who never made a mistake, never made a discovery.”

Samuel Smiles

That’s the beauty of science.

 

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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.

 

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Antibiotic resistance and what we can do about it

Something that has really struck me this year is learning about antibiotic resistance. It is such a widespread issue that affects every single one of us.

So, let’s start at the beginning…

What are antibiotics?

 

Antibiotics are widely used in the western world and most of us will have used antibiotics at some point in our lives. Antibiotics are great for many reasons, they are both convenient and effective. They prevent and treat bacterial infections by killing bacteria and can be taken in 5-7 day courses [1]. Before the times where antibiotics were widely used, people could die from something as small as a cut getting a bacterial infection. So, if they’re so easy to use and great at their job then what’s the problem?

What is antibiotic resistance?

Antibiotic resistance is the growing problem of bacteria becoming resistant to antibiotics. This happens because bacteria generally have a fast generation time, meaning the time it takes to asexually reproduce another generation is a short amount of time. Due to the fast generation time and large amount of offspring, mutations happen often. Sometimes the mutations may make the bacteria resistant to the antibiotics and therefore they will survive and continue to reproduce making a whole lot of antibiotic resistant bacteria. These bacteria then can’t be killed by such antibiotics.

This is a massive problem because antibiotics are vital in treating infectious diseases such as tuberculosis, pneumonia and blood poisoning [1]. We need antibiotics.

Antibiotic resistance is a widespread problem affecting us all but what can we do about it?

The overuse and misuse of antibiotics

antibioticresistnace

Antibiotic Costume“, Beatrice the Biologist (2014)

One of the reasons antibiotic resistance is on the rise is the overuse of antibiotics. Antibiotics kill bacteria and therefore should only be used to treat bacterial infections. In 2015 the World Health Organisation estimated that in half of antibiotic prescriptions the conditions are caused by viruses [2]. They do not cure viruses such as colds and the flu [3]. When people go to the doctor they often expect antibiotics even when they don’t need them. This is a problem because it exposes bacteria to antibiotics and therefore more bacteria become resistant.

There are things we can all do to prevent this such as not requesting or taking antibiotics when you do not need them, for example for a virus [2, 3]. Also make sure you finish the full course of antibiotics as otherwise some bacteria may survive and return.

Antibiotics in Agriculture

Antibiotics are commonly used in agriculture. They have many uses such as treating, controlling and preventing diseases [4]. They may also be used to promote animal growth [4]. There has been some evidence that antibiotic use in agriculture can impact antibiotic resistance in humans [4]. The use in animal growth is not necessary to the health of the animal and therefore is not necessary and may be contributing to antibiotic resistance. It is important for avoiding antibiotic resistance that we stop this from happening.

In New Zealand, so far there has been no evidence that in-feed antibiotics cause antibiotic resistance in humans [5]. However, there has not been much research. The Ministry for Primary Industries has said that most antibiotics in New Zealand agriculture can only be used to treat an individual showing symptoms or an individual in a group with others showing symptoms [5]. However, this ruling does not cover all the antibiotics and there is little information on exactly what is being done. We should be able to find out this information as it impacts us all. We should advocate for a decrease in the use of antibiotics in agriculture in New Zealand and across the world, especially as antibiotic resistance continues to increase.

Alternatives to antibiotics – phages

phageinfecting

Virus Nope“, Beatrice the Biologist (2015)

A phage (bacteriophage) is a virus that infects bacteria. So, like bacteria they can be used to treat and prevent bacterial infections. To kill bacteria, they enter the bacterial cell then replicate to make a large amount of copies then burst the cell by releasing all the copies.

Phage therapy is the use of phages to kill specific bacteria in our bodies and is an alternative to antibiotics. This can be done by people taking cocktails containing the phage that will target the host bacterium we wish to kill [6]. This was done and is still practiced in some places in Russia and Eastern Europe. However, it is relatively new in the western world.

An example of a potential use is in the disease Tuberculosis (TB). It is in the top 10 causes of death in the world, having caused 1.8 million deaths in 2015 and infecting around a third of the population with latent TB [7]. The overuse of antibiotics has accelerated the evolution of Mycobacterium tuberculosis becoming antibiotic resistant [8]. If we could find a phage that could infect this bacterium we could have a potential cure for TB.

This year we have been finding phages that can infect the bacterium Mycobacterium smegmatis. This is similar to the TB bacterium and therefore there is potential that some phages may be able to infect both. Hence, we could be contributing to potentially finding a phage that could infect TB and we are definitely adding to the scientific pool of data about phages which will help us discover more about this very important field in the future. In my next blog post I’ll be discussing this journey and the phage that I found.

References:

  1. World Health Organisation Antibiotic Resistance. 2016; Available from: http://www.who.int/mediacentre/factsheets/antibiotic-resistance/en/.
  2. World Health Organisation How to stop antibiotic resistance? Here’s a WHO prescription. 2015; Available from: http://www.who.int/mediacentre/commentaries/stop-antibiotic-resistance/en/.
  3. Antibiotic Resistance. Available from: http://www.health.govt.nz/your-health/conditions-and-treatments/treatments-and-surgery/medications/antibiotic-resistance.
  4. Timothy F. Landers, R., CNP,, et al., A Review of Antibiotic Use in Food Animals: Perspective, Policy, and Potential. Public Health Reports, 2012. 127.
  5. Industries, M.f.P. Antibiotics and resistance. 2017; Available from: https://www.mpi.govt.nz/food-safety/whats-in-our-food/chemicals-and-food/agricultural-compounds-and-residues/antibiotics-and-resistance/.
  6. Benjamin K Chan, S.T.A.C.L.-C., Phage cocktails and the future of phage therapy. Future Microbiology, 2013. 8(6): p. 769–783.
  1. World Health Organisation Tuberculosis. 2017; Available from: http://www.who.int/mediacentre/factsheets/fs104/en/.
  2. Nguyen, L., Antibiotic resistance mechanisms in M. tuberculosis: an update. Arch Toxicol, 2016. 90(7): p. 1585-604.
  3. http://www.beatricebiologist.com/
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What you Might Find in Your Garden

Stephen Fry: where would be the best place to discover an entirely new species?

Alan Davies: the Amazon rainforest (klaxon)

No as Stephen Fry pointed out the Amazon rainforest is not the best place to discover a new species. That honour belongs to your own garden (anyone’s garden that is).  The diversity of life in an ordinary suburban garden is such that  Jennifer Owen in 1971 did a study of the insects in her garden in Leicester and found 533 different species of wasps 15 of which had never been recorded in Britain before and 4 of which were completely new to science.

Wasps though are just multicellular life.  To give you an idea of how unknown bacteria and other unicellular life forms are 2 Norwegian scientists Jostein Goksoyr and Vigdis Torsvik  took a gram of soil from a forest near their lab in Bergen examined it very thoroughly and found 4-5 thousand new species of bacteria then they proceeded to do the same with a sample from a coastal area a few km away and found 4-5 thousand different species of bacteria.

However participants in the SEA phages programme search for something else altogether the even smaller viruses infecting these bacteria.  A virus is a strange entity in essence a piece of DNA surrounded by a protein which can attach to  a cell and “reprogram” it to produce more viruses. It is so small that even with most microscopes you won’t be able to see one. The  Scale of the Universe website can give you an idea just how small you should check it out it has a lot of interesting things.  What I might find just as interesting is whether a virus is actually alive…  that discussion can wait for another blog though.

Even if a virus is not alive you may be surprised to learn that it can be very useful and not just for terrorists intending on making an epidemic like you might see in an action movie. Phage therapy is the practise of treating a bacterial infection with a bacteriophage (virus) that will infect the bacteria. It may be another surprise to know that bacteria can themselves get sick (I don’t think I ever even considered the idea before reading about it) but it does happen.

In any case phage therapy has been suggested as a way of treating bacterial infections especially those that have become immune to antibiotics. In fact this form of treatment has already been used in parts of Eastern Europe since the 1940s. Several universities include a course discovering and studying bacteriophages. Massey University, Albany campus is one of these. The phages discovered by their class this year may one day provide a preventative treatment for tuberculosis or leprosy ( although the later is thought of as a thing of the past it still does exist).Image result for searching for a new species

Most of these are found in the gardens of scientists and other equally mundane locations. So if you don’t think there is anything interesting in your garden then I think maybe you should look harder.

 

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Te Ao o ngā Huakita me ngā Huaketo: The world of Bacteria and Viruses

Tēnā koutou katoa.  Ngā mihi nui ki a koutou, ki ngā manuhiri o taku pae tukutuku.

Welcome everyone to my blog.


Those of you who know me, will know that science is my niche and I often get very excited about new developments and technologies that arise.  This time however, I am excited because I have the opportunity to share with you, my whānau and friends, a glimpse into a world unseen.  A world that existed long before you and I, long before Māui pulled Aotearoa from the ocean, even before time its self.  The world of bacteria and viruses!

Te wero – your challenge; no matter your age or background, is to step out of your comfort zone and aim to learn something new.

If you went onto Google right now and searched ‘why are bacteria…’ the first thing that Google suggests is ‘why are bacteria important’.  Most of the time, we only hear about bacteria when someone is sick; so how come scientists are always going on about how cool and important they are?

I have a well-known adventurer here to join us.  Together we will explore the world of bacteria and viruses, so that we can understand why they are worth knowing about.

Māui! Nau mai haere mai – Welcome!

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The demi-god, Māui-tikitiki.

Haere mai – come on, we are going to explore the world of bacteria and viruses!  Our first stop, OURSELVES.

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 Yes we do!

We have just as many bacterial cells on the inside and the outside of our bodies, as we do human cells (4).  But don’t freak out!  Most of these bacteria are friendly.  They help  us  digest our food, improve our immune system and help fight off bad bacteria!

Each of you have a different mixture of bacteria which depends on your genetics, your environments, your diet and even whether you were born by C-section or not.

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Every individual has a different mixture of bacteria that make up their microbiome.

This environment of bacteria is called our Microbiome and they play a very important role in our health.  We need to make sure we are taking care of our bacteria, because they take care of us (2).

You might be asking yourself, how do I take care of my friendly bacteria?

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Our microbiome helps fight off bad bugs.

Great question! Well I’m no doctor, but what I do know is that bacteria LOVE food that is full of fiber.  Things like fruit, veges, nuts and legumes.  Our microbiome break this fiber down into useful molecules that we can absorb.  For example, small chain fatty acids that line our guts and act as a safety barrier.  Small chain fatty acids also assist our immune function and reduce inflammation. (2)

Therefore, the more fiber-full-food you eat, the more friendly-fiber-digesting bacteria you will have.

If you are eating low fiber, high processed food all the time (junk food), you are starving your friendly bacteria!  5 plus a day doesn’t JUST keep the doctor away, it also keeps our friends alive.  Let’s take care of our bacteria whānau.  After all it’s the one culture we ALL have.

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Take care of your culture. (Ravel et al, 2017)

#feedthefriendlies

To watch a video about this click here!

Now, we did mention before that there are also bad bugs.  These few bad apples can cause us to become very sick.  However, since antibiotics were discovered in 1928, previously deadly infections like pneumonia or TB are now easily curable!

Antibiotics are naturally occurring or synthetically produced chemicals which prevent the growth of bacteria.  They are our unseen super heroes!

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Yes Māui, like you (Wu, et al. 2014)

Our next stop is Te Ao Tūroa – the natural world.

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Bacteria are found everywhere in our natural environment; they assist natural processes just like how they assist us.  They can be found in the ocean, in our forests, and on our crops and livestock.

However, humans are not the only ones that suffer from the wrath of certain bad bugs.  There are bacteria that infect cows, horses, cats, dogs, native plants and crops.  These are important things to consider, especially in New Zealand where so many of our people rely on agriculture and horticulture.

Additionally as the kaitiaki – guardians of Te Ao Tūroa, it is our responsibility to understand all the threats to our environment, the seen and unseen.  Aotearoa is one of the strictest countries in the world when it comes to biosecurity.

Our health, jobs and even security can be affected by bacteria.

Whakarongo – Listen up!

Whānau, there is an important issue we need to be aware of in Te Ao Huakite – world of bacteria; and that is antibiotic resistance.

Antibiotic resistance is the ability for certain bad bacteria to withstand the effects of antibiotics.  This is very serious issue as there are few alternative methods to effectively fight off these bad bugs!

So how do these bad bugs become resistant?

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Antibiotic resistant super bugs (Wu, et al. 2014)

Over many generations, processes like natural selection mean that each generation of bacteria gains random mutations (for further explanation).  The effects of these mutations on the bacteria can range from harmful to no effect at all; but on the rare occasion, one of these random mutations will allow that bacteria to be antibiotic resistant!  Like gaining an extra super power.

This creates a strain of antibiotic resistant bacteria, that are called SUPER BUGS.

An example of a Super Bug is Methicillin-resistant Staphylococcus aureus or MRSA.  Staphylococcus aureus is a bad bug that can cause skin infections, pneumonia and sepsis.  The Super bad bug MRSA has gained random mutations that make it resistant to the previously effective antibiotic called, methicillin (6).

MRSA is a problem in Aotearoa. 

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Staphylococcus aureus mutates into MRSA (Wu et al, 2014)

There have been 8 different strains of MRSA found in Aotearoa.  In 2017, a total of 740 people were infected with one of these strains.  The people who were most likely to be infected were young people, and people of Māori and Pasifika decent (3).

It is currently estimated that antibiotic resistance accounts for 700,000 deaths worldwide.  It is predicted that by 2050 this number may rise to 10 MILLION deaths (1).

Kāore pai – not okay. 

So, if the Super Bugs can defend themselves against the antibiotics, then how do we fight them?  Here are a few things we can do to address this issue (6);

  1. Stop using antibiotics to treat infections that can heal them selves
  2. Allow naturally occurring bacteria to hang around, as competition for the bad bugs
  3. Find alternative ways to fight the bad bugs

One of the alternative methods to fighting bad bugs is using Bacteriophages (phages).  They are a type of virus that infect bacteria.

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Bacteriophages are a type of virus that infect bacteria

There are 1031 bacteriophages in the world.

That is 10,000,000,000,000,000,000000,000,000,000 phages!

He maha – a lot!  Each type of phage is able to infect and kill one type of host bacteria.
This makes them highly specific, unlike antibiotics.  In countries like Russia, Poland and Georgia, they sometimes still use phages to treat bacterial infections rather than antibiotics (5).

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Māui sharing his insightful thoughts

Yes there is Māui.  They have a huge potential to influence our day to day lives as human beings, which is why everyone should take time to understand their world.  Especially with problems like antibiotic resistance and new, exciting discoveries with bacteriophages.

Nō reira – Therefore

Ngā mihi whānau for your time and ngā mihi Maui for coming along.

I hope you all have learnt something useful, that you can pass on to your friends and whānau.

Next time we are going to investigate our phage friends; to understand exactly what they are and how they can be used to solve problems.

Stay tuned for the next adventure into the world unseen, Te Ao o ngā Huakita me ngā Huaketo.

Ka kite anō!

Anezka Hoskin

References

  1. O’Neill, J. (2016). Tackling drug-resistant infections globally: final report and recommendations. The review on antimicrobial resistance.
  2. Ravella, S. (Educator), Foerster, A. (Director, Animator, and Story board Artist), Nacamulli, M. (Script Editor) & Turner, J. ( Art Director, Designer, Illustrator and Character Designer), (2017).  How the food you eat affects your gut: TED Ed Lessons Worth Sharing.
  3. Richardson, A., Desai, U., Mowat, E., et al (2010). Annual survey of methicillin-resistant Staphylococcus aureus (MRSA). Retrieved from: www.surv.esr.cri.nz
  4. Sender, R., Fuchs, S., & Milo, R. (2016). Are we really vastly outnumbered? Revisiting the ratio of bacterial to host cells in humans. Cell, 164(3), 337-340.
  5. Sulakvelidze, A., Alavidze, Z., & Morris, J. G. (2001). Bacteriophage therapy. Antimicrobial agents and chemotherapy, 45(3), 649-659.
  6. Wu, K. (Educator), Underhill, B. (Animator), & Gendler, A. (Script Editor), (2014).  What causes antibiotic resistance?: TED Ed Lessons Worth Sharing.  

Want to learn more about antibiotic resistance? Click here

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The Pioneers of Phage Virology

One of the most surprising things for me in starting my ‘phage journey’ was just how long we have known about bacteriophages and their potential.  And yet we still have so much to learn!  It made me curious as to who discovered phages in the first place, and how.  In reading the blog posts from previous Massey University Phage Hunters I really enjoyed the variety of content, especially posts that explored a particular area of interest for that student. So, I decided to look into the history of phage discovery, and add my ‘slice to the pie’.  I hope you enjoy it.

Todar - Electron Micrograph (left) and model (right) of bacteriophage T4

Electron micrograph (left) and model (right) of bacteriophage T4 (Todar, 2012)

Ernest Hanbury Hankin was a bacteriologist working in India, who in 1896 discovered that cholera bacteria were susceptible to an organism in the Ganga and Yamuna rivers (Wittebole, Roock, & Opal, 2014).  The nature of the organism was unknown, but it was small enough to be able to permeate millipore filters used to retain bacteria in solution.  He didn’t know what it was, or how to see it, but he knew it was there.

In 1915, Frederick William Twort was trying to grow a viral strain on agar when he observed not only colonies resulting from bacterial contamination of his sample, but a strange morphology of what he assessed to be colonies of different bacterial strains (Kutter & Sulakvelidze, 2004).  They appeared either entirely or semi-transparent, and Twort realised that these ‘colonies’ were actually areas of bacterial cell degeneration.  Twort hypothesised that this could be due to an enzyme or virus too small to be seen by microscope.

Now remember, every time one of your fellow students is disappointed at returning to the lab and finding no plaques, despite the care and consideration that went into their method, you can tell them that over 100 years ago, ol’ Freddy got them without even trying.  Not only that, but they were the result of what would today be considered poor lab technique.  Some people get all the luck.

Stent - A picaresque genius

Felix d’Herelle (Stent, 2000)

Two years after Twort’s happy accident, microbiologist Felix d’Herelle made the independent discovery of a microorganism which appeared to utilise and destroy bacteria in the process of its own reproduction, resulting in cleared areas of cell death on bacterial lawns.  He called the cleared areas ‘plaques’, and concluded that they were the result of a virus that attacked bacteria (Kutter & Sulakvelidze, 2004).  These characteristics were the basis for d’Herelle naming this type of virus Bacteriophage.

I tried to figure out how d’Herelle knew that these strange little bacteria-assassins were viruses, but access to a published copy of his work that is accessible either online or in little ol’ New Zealand, and also conveniently published in English, may take a little more digging to come by.  I don’t think it’s unreasonable to assume that viruses were at the forefront of every microbiologist’s mind back then, having been only recently discovered in the late 1890’s (Zimmer, 2015).  I try to imagine what their thought process would have been to reach the conclusions that they did.  I suppose that despite a lack of knowledge, it may have been an easier process to discover new evidence, rather than have a wealth of education as a resource to search for the answer to a specific question.

The concept of ‘phage therapy’ was also introduced by d’Herelle (Fruciano & Bourne, 2007); he was one of several microbiologists who worked to isolate bacteriophages that targeted several known bacterial pathogens, and began to experiment in treating disease with phage therapy.  These treatments were successfully involved in the combat of multiple staphlycoccal, intestinal and systemic infections.

Lipman - vials of phage from Georgia, Eurasia

Vials of phage from Georgia, Eurasia (Lipman & Ferguson, 2015)

Phage therapy was not widely embraced by Western countries, though it is practiced extensively in Russia, Poland and Georgia as a result of previously restricted access to antibiotics (Reardon, 2014).  Its relative safety and predicted longevity in comparison to antibiotic treatments have many touting it as the way forward in a time when the growing antibiotic resistance of many bacteria is a rising threat.  If this is where we stand after 100 years of limited bacteriophage research, imagine what we could potentially achieve in the future with more widespread involvement.

 

References:

Fruciano, D. E., & Bourne, S. (2007). Phage as an antimicrobial agent: d’Herelle’s heretical theories and their role in the decline of phage prophylaxis in the West. The Canadian Journal of Infectious Diseases & Medical Microbiology, 18(1), 19–26.

Kutter, E., & Sulakvelidze, A. (Eds.) (2004). Bacteriophages: Biology and Applications. Florida: CRC Press.

Lipman, M., & Ferguson, A. (2015). Bacteriophage: Good Guys of the Virus World. Evergreen Magazine, Spring/Summer.

Reardon, S. (2014). Phage therapy gets revitalized. Nature, 510, 15-16.

Stent, G. S. (2000). A picaresque genius. Nature, 403, 827-828.

Todar, K. (2012). Bacteriophage. Todar’s Online Textbook of Bacteriology.  Retrieved from http://textbookofbacteriology.net/phage.html

Wittebole, X., Roock, S. D., & Opal, S. M. (2014). A historical overview of bacteriophage therapy as an alternative to antibiotics for the treatment of bacterial pathogens. Virulence, 5(1), 226-235. doi:10.4161/viru.25991

Zimmer, C. (2015). A Planet of Viruses: Second Edition. Chicago: The University of Chicago Press.

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The “bomb blast” and the “sneaky ninja”: Antibiotics vs Phage Therapy

As suggested by the title, these two different methods for the treatment of bacterial infections could not be more different. While antibiotics operate by ridding the body of any bacteria they may come across (good or bad), phage therapy is a process that targets just the infection-causing “bad” bacteria. Antibiotics are therefore reminiscent of a large bomb blast- there is always a specific target to take out, but the casualties are spread far and wide. Phage therapy on the other hand can be thought of as the process of employing little ninjas to sneak in and rid the body of the nasty infection-causing bacterial cells, with no other cells being affected.

But first, antibiotics:

Antibiotics are medicines, either naturally occurring or semi-synthetic, which inhibit the growth of microorganisms, specifically bacteria, thus allowing the antibodies of the host to kill off the infection-causing bacterial cells. Antibiotics are administered in liquid form, as tablets, in the form of creams and even through IV (intravenous) therapy in the case of more extreme infections. The problem with antibiotics is that they are not able to tell the difference between “good” and “bad” bacterial cells, and so they end up affecting the growth of important bacteria, such as those in the gut of mammals, and hence probiotics must often be taken simultaneously.

From what I have read on various websites, I think that it would be quite accurate to assume that most people in the Western world have either heard of antibiotics or have taken antibiotics or both. The discovery of the very first antibiotic, Penicillin, happened in 1928 and was discovered by Alexander Fleming. After many years of research, in 1943 Penicillin became available for widespread production and clinical use. Other antibiotic discoveries followed in 1943, 1947 and 1952, the Golden Age of antibiotic discovery in the 1940’s to 1950’s seeing many new natural antibiotics being isolated from soils. However in 1948 Penicillin-resistant strains of Staphylococcus aureus had already been circulating and a global pandemic followed between the years of 1948-1962. This should have been a clear warning of the dangers of the ever-evolving tendencies of bacteria. But, in 1960, Methicillin was introduced for chemical use- the first semi-synthetic penicillin derivative. By 1962, just 2 years later, there were already clinical strains of methicillin-resistant Staphylococcus aureus appearing. There is an interesting timeline here that extends on the topic of antibiotic discovery, but a common trend you will see is that many bacterial strains develop antibiotic resistance, and this is a serious problem.

There was recently an article in the Washington Post, detailing the arrival of the “superbug” in the U.S., a strain of E.coli  (seen in the image below) found to be resistant to the antibiotic colistin. This antibiotic has been known as a “last-resort” drug, that can have serious adverse side effects and which is usually administered in the case of especially tenacious bacterial infections resulting from multi-drug resistant bacteria. The deaths associated with multi-drug resistant bacterial infections is on the rise, and As more drug-resistant strains of bacteria emerge, we get closer and closer to the end of the antibiotic-era, and the exploration of alternative treatments is becoming essential.

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The reason why these drug-resistant strains emerge in the first place, is usually as a result of two scenarios: one, the antibiotics are prescribed long before they are necessary, in a stage where the antibodies of the host could still fight off the infection Or two, the antibiotics are not taken correctly- that is, people stop taking the antibiotic when the infection appears to have subsided. Either way, the bacterial cells are provided with the opportunity to become resistant when we don’t succeed in killing off every individual cell. And given the rapid growth rate of bacterial colonies, it doesn’t take long before the surviving individuals multiply, passing on the drug-resistance gene to their offspring.

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Now, Phage Therapy:

Given the situation we find ourselves in, exploration of alternative treatments is of great importance. And to give you a bit more of an introduction to the ideas behind the existing alternative to antibiotics, watch this clip (by our very own Dr Heather Hendrickson!) about the “Microbial ninjas” we call bacteriophages, just like the one seen in the image above.

Phage therapy was widely used in the United States, former Soviet Union and Western Europe in the 1920’s and 1930’s, until the discovery of antibiotics resulted in the abandonment of phage research in the U.S and Western Europe. In the countries of the former Soviet Union, phage therapy and research is ongoing. Most notably in the country of Georgia, where the Phage Therapy Centre is found in Tbilisi (see the map below).

The link provided will take you straight to their website, which has LOTS of in-depth information regarding phage therapy practices, so take a look if you like.

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The beauty of phage is that they are specific to the bacteria they infect, and so they will not harm the “good bacteria” in the host body suffering from a bacterial infection. It is of course by chance that phage infecting a specific bacteria are found, but that is why continuous research is necessary, and also so that in the case of bacteria developing resistance to one phage, another may be ready and waiting to take its place.  And due to the estimated abundance of phage on Earth- about 10^31 phage- the chances of finding more than one phage infecting the same bacteria are quite high. The phage are of course naturally occurring, and so are much cheaper to “produce” in large quantities than antibiotics.

Although it is necessary to isolate and purify phage before administration, there are much-researched and straight forward ways of doing this. The purified phage with a specific, known titer can be administered through liquid preparations, tablets and also in the form of dressings for infected wounds, and may be given in conjunction with antibiotics if the infection is particularly persistent. The most successful use of phage is in the treatment of gastrointestinal infections and those infections associated with open wounds or burns.

In summary, these sneaky ninjas called phages have a lot of potential for success, as has been proved by the treatment carried out in Georgia for all of these years. This so-called “forgotten medicine” has all the makings of a successful alternative to antibiotics, although it may need to be more rigorously studied here in New Zealand before it is recognised as safe to use on people and animals to treat infections.

To conclude this blog post, I’d like to introduce you to my phage, Phage PlainJane. This link has all of the details outlining where this phage was captured, tamed and finally dissected to get a better look at it, as found on the PhagesDB.org website. From the EM image, she may just appear to be a regular siphoviridae phage, as is common in phage isolated using Mycobacterium smegmatis, but who knows what’s on the inside? Potential can often be found in the most unexpected places. And regardless of the whether or not PlainJane is something special scientifically speaking, “she” will always be special to me- a symbol of this extremely rewarding and oddly fulfilling process of finding and naming a new, unique organism (no matter how small).

It is now an experience that has been ticked off, before it had even appeared on my bucket list!

√ Discover a new, unique (tiny, ninja) organism.

Goodluck to all the future phage-hunters, I hope you enjoy the journey as much as I did.

All the best,

Catherine 🙂

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Giving Life a Chance…

Urinary  tract infection during pregnancy using public facility and I also provided some information on how to dangerous it can be for both, unborn child and expectant mother. Now I would like to have a quick review of a treatment options  available  in eastern Europe.

So why not treat infection early before it progressed to advanced stage and cause so much irreparable damage?

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Sadly enough, but most of the treatment options available now days in Western countries are limited to antibiotics only, regardless of  numerous facts that confirm the negative impact of administering  antibiotics during gestation. Beyond doubt, using antibiotics during pregnancy is not recommended as it has teratogenic effect on unborn baby 1 2.   Teratogens are substance or organisms cause abnormal foetal development and birth defects.

Under normal conditions medication metabolites are processed in the liver and excreted by the kidneys with urine. When kidneys are damaged the rate of excretion is reduced critically, which leads to prolong circulation of drug in a blood. Administration of antibiotics has to be done with the great caution as even slightly increased dosage can exceed the ‘safe’ level in the blood and even lead to renal failure 3.  Even though drug toxicity increases dramatically as result of malfunctioned kidneys that are unable to completely get rid of wastes, serious infections like pyelonephritis still require prolong treatment with large amount and different combinations of antibiotics.

Apart from adding stress to earlier impaired kidneys, continuous drug administration often causes bacteria to acquire resistance to certain antibiotics, which makes therapy not only risky but also ineffective.

Once bacteria acquire resistance to particular class of antibiotics designed to target specific group of pathogens, the risk of losing control over the disease is high.  If no alternative treatment available the only option is either immediate pregnancy termination if foetus is under 23-24 weeks of gestation or labour induction if baby reached 24 weeks (25 in some countries).  Starting labour artificially before 35 weeks of gestation (normal pregnancy lasts 40 weeks) can often have dangerous if not tragic consequences, especially when child’s growth was stunted by infection.

Due to toxic effect on child and limited therapeutic potency of antibiotics, eastern European medicine developed a new approach to treatment of UTI during pregnancy.   In former Soviet Union lytic bacteriophages are widely prescribed for large number of various infections.

Because of exclusively high specificity of bacteriophages in relation to host, the pathogen species have to be identified prior to start of the treatment.  Infections during pregnancy require urgent attention and due to time constraints treatment often starts before bacteria identified.  To avoid delays in therapy, Russian scientists manufactured the cocktail of assorted phages.  One of the most promising phage mixtures that target multiple strains at a time is sextaphage (pyobacteriophage).bakteriofag2 Sextaphage is a preparation of multiple phages against Staphylococcus, Streptococcus, Klebsiella, Proteus, Enterococcus, E. coli.  Phage preparation is regularly updated to meet current epidemiological requirements. Yet, it still consists of bacteriophages against most frequently isolated pathogens.

According to numerous clinical studies phage-treated patients had better pregnancy outcome with no registered cases of intrauterine transmission of infection or side effects if correct doses were administered.  In addition, phages can be used in conjunction with antibiotics, in order to reduce toxicity, but insure efficacy of complex therapy. Moreover, bacteriophages in a complex therapy greatly reducing the risk of sepsis causing by not properly treated infections in both mother in child.

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Making Humans. Troubleshooting & Tips. Part 1

A baby is God’s opinion that life should go on  (Carl Sanburg)

Childbirth is a life changing event for every couple. Although one may find expecting the new tiny family member fascinating and inspiring , I personally considered it somewhat tense and terrifying , mostly due to the tonnes of pregnancy precautions and widely accepted belief of inability to distinguish normal pregnancy symptoms from signs of conditions causing miscarriages, birth defects and stillbirth.

Meanwhile you can easily avoid being exposed to nasty chemicals  buying overpriced fruits, veges and shampoos  in organic stores ,  no one can guarantee that your environment or places you go are 100% microbe free.  Needless to say public toilets all of a sudden become your most visited place and, yes, cleaners do not check it right before you go in.

On a daily basis ‘normal’ immune system of a healthy person can easily handle frequent trips to wash room,  providing the fact you’re not a fan of adventure tours in Cambodia or volunteering in a rural Tanzania and by ‘toilet facility’ you assume a nice comfy semi-sterile room where you do not want to hold your breath like deep-sea free-diver . But if you are an extreme tourism lover, you’d probably need to read this article  as well, after all lack of adequate sanitation in some third world countries indicates that you have a great chance to get some mean exotic bug , even if you do not even plan to conceive.

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Thanks to western society with its modern hygienic standards that prevent spread of pathogens and disease they’re causing in healthy individuals, but remain ineffective for pregnant women which are still at high risk of getting bacterial infection.

I’d like to go into more details about Urinary Tract Infections since it is the most common type of infection women can get during gestational period.

Urinary Tract Infections, acronym UTI, is one of the major concern during pregnancy and one of the most common reason of miscarriage, foetal abnormality, severe prematurity and stillbirth.

During gestational period women are more prone to UTI, due to immune suppression and decrease in muscle tone of urinary tract.  A bacterium that reside in a bladder or lower urinary tract is capable to reach up the kidneys causing very serious medical complication called pyelonephritis.

When woman is pregnant, kidneys have to work to its full potential, since they are working for two, or even three people, clearing up wastes from mother’s body. Needless to say the amount of waste usually doubled or tripled during that period.  Already overloaded by extra work, kidneys can’t afford loosing even a tiny bit of its ‘productivity’ without compromising a well-being of the mother and foetus.

As it was already mentioned, the chances women to get kidney infection skyrocket at the time of pregnancy 1 2. There are number of factors influencing the susceptibility to infections during gestational period:

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  1. Increased production of hormone progesterone affects muscle tone and slows down the drainage of urine, causing it remain stagnant in a system followed by backflow of the bladder content into the kidneys. Bacteria settled in the lower urinary tract can easily access kidneys, instead of being washed out of the system 5 6 7.
  2. The systemic immunity of expectant mother is not yet well researched, but most scientific studies refer to it as being ‘weakened’ or ‘compromised” even though the main goal of immune system of a mother is to protect future offspring’s 8 9 10 11 12 13 14.

 

group-b-strep-infection-newborns792-x-612-131-kb-jpeg-xMeanwhile, instead of common immune response of an organism to urinary tract infections, such as fever and burning pain, the immune response of pregnant woman is rather unique. Atypical symptoms are always the huge impediment to the identification of the disease, due to similarity to common pregnancy symptoms.  Inability to detect pathogen’s invasion at an early phase often causes disease progression into advanced stage very rapidly 4.

The first symptoms to appear are severe nausea, vomiting, appetite loss and severe hypertension. Symptoms themselves require quick attention since they can lead to malnutrition, hypoglycaemia and dehydration.

A-premature-baby-undergoing-treatmentAs the pressure on already exhausted kidneys increases the chances of having severe birth complications become more and more possible.  If not treated early, pyelonephritis often result in foetal death, pregnancy loss, perinatal death and neonatal death. Perinatal death is when baby dies at late pregnancy, when she or he is just about to be born. Neonatal death is the death of live born child within first 27 days of life. The death occurs due to serious birth complications, prematurity or renal failure caused by infection. In cases when intrauterine transmission of infection occurs the child is already born with severely impaired  kidney and even if treated early will suffer from kidney malfunction  throughout his/her lifetime. Profound mental and growth retardation are also among of the most frequent consequences of pyelonephritis.

My next blog will be about treatment options available overseas.

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Phages: Nature’s (very) tiny demolition teams

 

Okay, so as a result of me being the procrastinator that I am, everything that needs doing gets done in the mystical land of “tomorrow”. And so here we are, approximately eight months from the time our Phage Hunt at Massey University started (in July 2015)- and this is my very first blog post. It’s been on the “to-do” list for a while now, about eight months actually, and so we have already concluded our Phage-Hunting labs. The next step will be the genome annotation of the bacteriophages that were found by our class of 2015/2016, and this intriguing process will begin in about four weeks’ time.

To back-track a little bit, the existence of the wonderful world of bacteriophages was first brought to my attention in a Biology of Cells lecture at Massey University in 2014 which was given by Dr Heather Hendrickson. The idea of such tiny organisms in such a high abundance around the world was almost impossible to fathom at that point. And to be honest, it still is, even though I have seen the presence of phage while working in the lab and have even seen them “in real life” using an Electron Microscope.

For those people who haven’t come across Phages before, and for those people who need to refresh their memories because it’s been a while (erhm, me), here is a bit of an overview of the world of phages:

Diagram for Blog Post 1

As illustrated above, phages are a specific class of viruses and they infect only bacteria. To give you an idea about relative sizes, a bacterial cell can be anywhere between 1-5µm across, this is x10-6m which is incredibly tiny in relation to us. Phages then, being parasites of bacteria, are even smaller than this, about 100-200nm across, which is 0.1-0.2µm which is x10-9m. There is an interactive video here that might help put things into perspective, but do you understand now why I can’t always get my head around their existence? In terms of abundance, it is estimated that there is a total of about 1031 phages on Earth. That’s crazy right?

While the bacterial cells can be viewed under a microscope, bacteriophages are far too small for this. Instead, we observe the presence of bacteriophages by plating our chosen bacteria- in this case Mycobacterium smegmatis (the not-so-nasty cousin of the well-known Mycobacterium tuberculosis) which has been infected with phage. For those not so familiar with scientific processes, basically we use those stereotypical petri dishes along with a growth medium to cultivate the infected bacteria. The bacteriophages then make themselves known by producing ‘plaques’. These are holes, either perfectly clear or cloudy, in the so-called ‘bacterial lawn’ that we have grown. They are the spots where the phage has ‘eaten’ away the bacteria. The images below are an example of this, there are both clear and cloudy plaques produced by the spot tests. See how the spots are not all the same? This indicates that there is more than one type of phage present- cool huh?

Just as plaque morphology varies, so too does the morphology of the phage themselves. However, regardless of their morphology, the general structure (basic components) of the phage does not change. The image below gives you a closer look at these important features. And here is a rather dramatic animation of how the phages ‘eat’ the bacterial cells to form these plaques. Remember that what you see is happening on a MUCH larger scale since the individual bacteria are so small and there are MANY of them, hence the plaques are visible to the naked eye.

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After being bombarded with all of that information, I suppose the obvious question now is- why bother studying these tiny masters of demolition? Of what use could they be to us?

To answer these questions I think it might be a good idea to talk more about bacteria and bacterial infections. So, first things first- what are bacteria?

Well, bacteria are prokaryotes or single-celled organisms. They are very diverse and are found pretty much anywhere on Earth that you could possibly think of. As with bacteriophages, bacteria have varied morphology but have basic components that are common to all bacterial cells, and the image below illustrates these. Bacteria are able to grow at exponential rates, meaning that entire bacterial populations increase in numbers at a very rapid rate when they are in their preferred environment. This becomes a serious problem when certain bacteria infect plants, animals or people, and in many cases antibiotics (which kill off bacterial cells) have to be administered.

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Diagram of a generic bacterial cell.

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As I mentioned earlier, we worked with Mycobacterium smegmatis in the lab, and this particular species is not pathogenic (disease-causing). However, it does belong to the genus Mycobacterium, which contains many known pathogens that are disease-causing in mammals, the most commonly known being Mycobacterium tuberculosis. M.tuberculosis is responsible for the deaths of millions of people around the world every year, and while these numbers have declined over recent years due to improved treatments and access to treatments, there is a constant threat of the TB bacterial cells developing a resistance to antibiotic treatments. (Click here to have a look at the WHO global tuberculosis report for 2015). Multidrug-resistant TB strains already exist and this is a cause for great concern, as it seems that new antibiotics are set to become a thing of the past (but more on this next time).

There is a solution to this problem though, and it’s called Phage Therapy. The basic idea is to use bacteriophages to treat bacterial infections, which would make perfect sense since their sole purpose is to find and ultimately destroy bacterial cells. It is a process that has been used at the Phage Therapy Centre in Georgia for the last 80 years, with varying degrees of success.

I intend to discuss this topic further in my next blog post, so keep an eye out for that in the next few days. But in the meantime, have a read of Sam’s story, a first-hand account of his experience with Phage Therapy- it’s very interesting indeed!

Until next time then,

Catherine 🙂

 

 

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