This is something I would like your input on:
How important is strain choice in virus/microbiology research?
Does it differ with certain aspects of research, like pathogenesis versus structural studies?
How far do we discount studies done on less 'wild-type' viruses?
How come people are happy working with these viruses?
We microbiologists uncover the secrets of how microbes interact with their hosts in order to better understand the infection process. We can use this knowledge to allow us to develop antiviral treatments, preventative vaccines and important applications for these molecular parasites, such as anti-cancer therapies.
All of this requires that we are studying what is real and what is actually going on in the world around us. This is all the more important when peoples lives depend on our work and when we are dependent on tax payers money to fund the research. Generally, this seems very obvious but you would be surprised about how much work is done on by far less-than-perfect viruses. This is because there a number of hurdles in the path of us understanding what is real-life.
Here's the problem:
Much of virology - and other sciences - use experimental models to understand what happens during a real-life infection. For example, much virus research is dependent on a mouse infection system where we inoculate the small mammal with large amounts of virus so that we can know what happens when a human comes into contact with the virus. This can obviously lead to a number of experimental artifacts. After all, mice are definitely not humans. And, so goes the old caveat: mice lie and monkeys exaggerate. This can also be extended to the avian influenza work that is recently in the news that documents the generation of a virus that can spread from one ferret (a good influenza model) to another. What happens in ferrets may not happen in humans.
Another example of this is from the point of the virus: not all viruses are alike. Many strains of a particular virus exist, some are naturally occurring variants that exist out there and are a result of normal evolutionary processes, while some have never existed outside of the virus lab. In general these viruses are very hard to get access to experimentally as the major method of virus isolation and growth is tissue culture and may itself alter how the virus behaves. This occurs through allowing the rapidly mutating virus to adapt to life in a test-tube, i.e to life in one particular cell type (which may not even be the natural host) and to life without a proper immune response. If we are keen to study something real we should therefore be very careful not to accidentally work on some laboratory curiosity.
To take one recent example (there are however, many, many more of these):
MicroRNA-221 Modulates RSV Replication in Human Bronchial Epithelium by Targeting NGF Expression
This paper, published a few weeks ago in PLoS ONE, seeks to understand how the human respiratory syncytial virus - an extremely important pathogen - affects the expression of specific small regulatory RNA molecules. They are particularly interested in how virus infection of the respiratory tract alters one specific protein involved in control of apoptosis. This work, funded in part by the US NIH may go on to form the basis of a pharmacological treatment to inhibit RSV infection and disease. It may also go on to influence a number of other groups researching the same area. This is generally a well done piece of work - and there is a lot of work done but the one problem of this (a major one in my mind) is what I talked about above, these guys used of a very bad experimental model of RSV.
This group - and many others - use a strain called RSV-A2, which is a well known laboratory adapted strain that has been grown too many times to count on cells that aren't natural (I think it was something like 52 replications on a cow cell line). This has resulted in a viral lab curiosity (see a differential pathogenesis paper here) yet it is still used in so much RSV research, even in a recent paper documenting the first non-human primate trials of an anti-RSV vaccine. This is all very important research which is neglecting real life. Although, much research is now being carried out exploring these differences and using more clinically-relevant viruses.
The same kind of problems plagued the measles virus field for years following the first identification of its receptor molecule, CD46. It turned out that after nearly a decade of research done, this receptor was only used by vaccine (lab-adapted) strains and not wild isolates.
What can we do about it:
This is only one example but I am sure that every kind of virus research is inflicted with these problems, where we can never be sure how to interpret results that are based on these viruses. Many times it is a trade-off over ease of experimenting versus applications of the results, as many times clinical isolates of viruses just do not grow as well or as fast as their lab-adapted cousins. In some fields of research, such as: structural work this is generally not much of a problem I don't think but in the likes of pathogenesis work where we rely on the 'real' virus we must concentrate on making sure what we are using in real.
To stem these problems we could at least back up your results with as clinically relevant (or if they don't cause disease, fully wild-type) viruses as possible. We could at most develop the reagents: a recombinant wild-type virus bearing the exact sequence of wild isolates or a collection of low passage viruses with same sequence as the wild viruses.
Othumpangat, S., Walton, C., & Piedimonte, G. (2012). MicroRNA-221 Modulates RSV Replication in Human Bronchial Epithelium by Targeting NGF Expression PLoS ONE, 7 (1) DOI: 10.1371/journal.pone.0030030
Some very good points raised.
ReplyDeleteLab adapted virus strains often lead to misleading data that poorly reflect the situation in vivo. This is particularly a problem when scientists don't realise the virus is adapted, or in some cases stick their head in the sand and ignore the fact.
However, almost all biological research involves compromise. For instance, primary isolates of hepatitis C virus (HCV) will not grow sufficiently well in tissue culture to allow robust study of the virus life cycle. This held back HCV research for years. But in ~2005 a virus taken from a patient in Japan with raging acute hepatitis turned out to replicate just fine in the lab.
By virtue of its ability to grow in vitro this virus cannot be considered normal. However, it opened the door to thousands of fantastic studies in to HCV and could eventually lead to new therapies.
Compromises will always be necessary. But by acknowledging the caveats and interpreting the data with care, valuable work can be done. Although, problems will continue to arise when scientists unwittingly or willingly ignore the intrinsic flaws in experimental systems.
Joe, thanks for your comments - I had in particular thought of the HCV situation.
ReplyDeleteBut I think this example is different from the others where groups do - as you say - stick their head in the sand and fail to discuss that their results are based on a lab adapted strain.
Maybe they don't realise that they are doing but I think it would be hard for any PI not to know what they are working with.
I think compromise is important but if you are a group leader wanting to work on a certain aspect of a virus, why not go and develop the proper tools to do so correctly? Without backing your work up with wild-type/clinical isolates I find it extremely difficult to interpret the work. More so with pathogenesis work that relies on every aspect of the viral life cycle.
Great post on a really important topic that no-one likes to talk about.
ReplyDeleteI would like to think that most scientists give some thought to what bacterial strains they use as their models, but I have been at a prize winning conference presentation where someone used DH5alpha e.coli for a mouse model of EHEC. For those of you who didn't immediately facepalm upon reading that, it's like using a toothless blind chihuahua to model a bear attack.
But there are a few arguments for using "lab adapted" strains of bacteria. Often lab adapted strains have publicly available genomes, and are more amenable to genetic manipulation. So they can be used to analyse specific metabolic or pathogenic processes.
There are some who I've worked with who take the view that "lab -adapted" strains are an excuse to not develop better techniques for working with clinical strains.
Working with strains that no-one else has worked on increases your chance of finding something no-one else has spotted before, and isn't that why we went into science in the first place ?
Defective Brain - it is a very important topic and sadly it isn't really talked about, officially at least. If you are doing tissue culture work with mammalian cells for example, quite recently people have begun to focus on primary cells but the same cannot be said about true 'wild-type' pathogens.
DeleteThe DH5alpha thing is hilarious but quite worrying how that can be done.
I agree whole-heartedly with your last two points, but sometimes when you have funding for three years it's not that easy. But maybe with all the sequencing advances etc this might become a non-problem in the next couple of years.
The utility of clinical strains needs to be weighed up depending on what you need from the model. In some cases it could well be worth it to bang your head against a brick wall of failure for 3 years until finally cracking the problem. In others, wasting time developing a new model when there are perfectly relevant ones already available may not be worth it.
DeleteTo be fair, most common lab strains did start out as clinical isolates. But then the world moves on, and the strains circulating in the community change, and they become irrelevant. Or they get passaged into impotence.
Equally, today's hot new clinical strain could be tomorrow's lab adapted strain. Working with clinical strains shortens the distance between the bench and the bedside. But it doesn't eradicate it.
There will still be biases based on what gets sent from the wards or the community, and what can grow in the lab. To some extent, there will always be a need to play catch up with microbes evolving in the community.
Sure, but I think that any work on pathogenesis in vivo needs to be as close as possible to what you actually observe. Especially if this is being used for the likes of antibiotics/vaccine trials. You could waste a lot of money on something that is not real. But of course for other subjects I don't think it's necessarily so important. Although, if you can develop a more wild-type model, why not?
DeleteI agree with the lab strains being clinical ones at some point in time. That's something you will just have to keep updating. As long as people are using a clinical one with low passage history then it should ultimately be fine. I think. It's just it seems some people are OK using a 50+ passaged microbe and they think what they see is real.