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Genetic barcoding and mythical creatures

10/12/2012

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As a fly enthusiast, I understand how daunting a task identifying species can be. The minute details, the crazy terms: it can all make you lose your head, especially when you’ve gathered a seemingly infinite amount of specimens. But, what’s a scientist to do? 

You could hunker down at a microscope and wait until your eyes cross, or you could head down the road of genetic barcoding. Now, simmer down, you taxonomists. I don’t plan to argue you guys out of your jobs. In fact, I have my own criticisms of barcoding, but just humor me for a moment.  

Genetic barcoding works by sequencing small DNA portions from unknown organisms and comparing those sequences to a barcode library. So say you’ve collected a bunch of something, let’s say unicorns from the North Pole as everyone knows all magical ponies live in the wintery north. Well, as a well-known unicorn scientist you are aware that there are several cryptic species of unicorns. This means that two or more species appear morphologically similar but, by at least one of the many species concepts, are still considered separate species. A quick PCR analysis, PCR gods forgiving, and a BLAST to the NCBI database could tell you which mythical unicorn species you now possess (should the barcode library of unicorns be complete).

Okay, I may have lied. Unicorns don’t really exist (outside the imagination of yours truly), but the problem of cryptic species does, along with a myriad of other identification issues such as morphological variation within species and even between adults and juveniles. Have you ever looked at drosophila larvae? They all look like squiggly, little, wormy things, every single one of them. Aside from some neat distinguishing behaviors –  a few fling themselves like trapeze artists – you couldn’t tell them apart.

So, it makes sense that a useful tool like barcoding has received so much attention, but let’s not get carried away. This isn’t the messiah come here to solve all our problems. The way I see it genetic barcoding is the microwave of the 1970’s housewife: a new tool for the modern taxonomist. It heats your food in mere minutes, but you can still burn the pot roast. Criticisms include incorrectly identified species sequences, a substantial error rate, and lowered ability to distinguish between recently diverged species. These comments all point towards the necessity of well-studied taxonomists to make final decisions.  

Me? I’m sticking to the microscope for now. Having a good grasp on taxonomic identification seems like it will always be a useful tool.

Jessica Craft
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DNA barcoding and gene-chips 

10/12/2012

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When I was 19 years old I visited the Organization of Tropical Studies’ La Selva Biological Station in Costa Rica. Upon a nature hike with a resident researcher, a hypothetical, nearly sci-fi idea was thrown out for ways to significantly improve field work. The scientist painted the picture of a futuristic pocket-sized chip that could puncture leaf or animal tissue, do a lightning fast DNA extraction and PCR, query a genetic database, and within minutes identify a specimen – right in the field! He proclaimed that this invention would allow scientists to categorize greater biodiversity, understand ecosystems more fully, and help to clarify the taxonomy and phylogeny of tropical species.  

Daniel Janzen, a renowned tropical ecologist and professor at the University of Pennsylvania, is a major proponent of this theoretical device. Janzen has been involved in the 'Consortium for the Barcode of Life’ project, which includes members such as the Natural History Museum in London, the Smithsonian in the US, the University of Guelph in Canada, Rockefeller University in New York, and a host of other institutions. The goal of this research consortium is to use a single DNA sequence, (cytochrome oxidase I, a mitochondrial gene), to essentially tag, or “barcorde” every species on earth. Having one gene with which to identify all biodiversity is a lofty task that will require many skilled technicians in functioning genetic labs, as well as taxonomic experts to assign appropriate names and voucher specimens to all of these sequences. Still Janzen suggests that with the use of the proposed ‘gene chip’ the process could be conducted by a “six-year kid walking down the street.”  

Progress has already been made in the construction and usage of this 'theoretical' device. Mesa Tech International has developed the ‘DNA dipstick,’ a hand-held, battery-powered, disposable device that can identify nucleic acid sequence-level data within hours. This device has been used to identify microbial pathogens in agricultural crops and animals and thus improve human health. DNA microarrays have also been used in the Fish&Chips project which hopes to identify and categorize marine biodiversity. This project uses a ‘bio-chip’ made of glass that contains oligionucleotides fixed to the chips’ surface, which acts as a probe to bind complementary target DNA sequences by hybridization. This group also has a Phytoplankton Chip and Invertebrate Chip. With such technological developments in recent years, the quick identification of specimens in the field, as proposed by the Costa Rican researcher some years ago, suggests that this goal is not so far-fetched. DNA barcoding and the use of gene-chips will undoubtedly herald science into a new era, as we begin to database and identify genes of all of earth’s species. 

 Iman Sylvain
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On barcoding, bandwagons and the nature of scientific revolutions

10/12/2012

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Several recent events have me reflecting on how science is done and how different schools of scientific thought turn over through time.  I'm teaching a grad class in phylogenetic methods for the first time since 2007 and I've noticed a big difference in the students.  In the past, their emphasis has been on understanding the nuts and bolts of how to generate phylogenies.  While the students this year are still interested in building trees, I'm getting the sense that they view tree building as a means to an end, rather than a valid activity in and of itself.  Student interest seems to have shifted more towards using trees to test evolutionary hypotheses.  While the sample size is small, this echoes what I've heard from colleagues about two courses taught in integrative biology, IB200A (phylogenetic reconstruction) and IB200B (phylogenetic hypothesis testing).  Enrollment in 200A is dropping relative to 200B.  

It's an interesting phenomenon and makes me think we may be in the midst of another paradigm shift (albeit a small one) in how systematics is done.  Looking back over the years, you can clearly see turnovers in schools of scientific inquiry.  Here's a short list:

Numerical Taxonomy
Starting in the early 1960s, numerical taxonomists brought a quantitative approach to taxonomy and systematics that previously been absent.  This was driven largely by statisticians (Sokal, Sneath) and the notion that careful measurements could lead to improved taxonomic hierarchies.  During the 1970s this field fractured into phenetics and cladistics.  Then the cladists ate all the pheneticists. 

Cladistics (and cladists)
While cladistics and cladists are tightly linked, not all people who practice cladistics are cladists and not all cladists always employ a strictly cladistic approach.  In essence, it's a semantic argument, something that all good cladists enjoy.  I define cladists as those followers of Willi Hennig who espouse a parsimony-only approach to systematics.  They aggressively routed the numerical taxonomists in the 70s and then stuck around to rail against likelihood, Bayesian analysis, and, in some cases, evolutionary inference itself.  For more detail on this era of systematics, check out the chapter in Joe Felsenstein's Inferring Phylogenies book.

Molecules vs. morphology
Starting in the mid-1980s, the introduction of PCR led to a technical revolution in systematics.  Suddenly, everyone was scrambling to sequence DNA in his or her favorite organism and use it to generate phylogenies.  Like most of the previous theoretical and technical advances in systematics, DNA promised to "fix everything."  This, of course, hasn't come to pass and, even now that we can sequence entire genomes, some systematic questions remain difficult to approach.  What did happen was a massive shift in resources, both in terms of grant funding and jobs offered, with the traditional morphologists being on the losing end of things.  This led to a lot of animosity - I can still remember being called a "moleculoid" by some of my older colleagues.  Luckily, this has largely blown over and most systematists take a holistic approach to understanding relationships in their focal taxa.

DNA barcoding
In some ways barcoding is a spin off of the molecules vs. morphology debate.  The notion here is that taxonomy isn't really needed now that we can use DNA sequence to uniquely identify (or barcode) species.  While DNA approaches are important techniques to have in your taxonomic toolkit, throwing out all by a single character system (the COI gene if you work on animals) in your taxonomy is ridiculous.  And many people have pointed this out before.  The initial DNA barcoding push was really more of a marketing campaign than a novel scientific approach and, once again, a more inclusive approach is being taken.   

Statistical phylogenetics
The idea that phylogenies are statistical statements about evolutionary history and can not only be viewed as hypotheses but also used to test hypotheses is the predominant paradigm in modern systematics.  More advanced analytical techniques, increased processor speed, and the introduction of model-based approaches have all helped shaped modern phylogeneic systematics.  Powerful statistical methods are currently causing an expansion of systematics and driving the "use of trees" over the "building of trees."  I think this is a normal, natural outgrowth of the field and will hopefully continue to drive it forward. 

My own work is moving away from tree building and more into the area of community assembly and interaction so I've been reading a lot about phylogenetic community ecology (PCE from here on out - too much to write) as a way to merge the fields.  I ran across this interesting blog post a few days back where the author, Jeremy Fox, makes the case that PCE is a "bandwagon." He makes bring up some good points (although he uses a pseudo-subjective literature review to do so) and the post is worth a read.

This all leaves me wondering, however, if there's really anything wrong with any one field or subfield jumping on a bandwagon.  This is, at least if you take an historical perspective, how science moves forward.  For example, organismal biology jumped hard on the DNA bandwagon in the late 1980s/early 1990s, eliminating entire -ologies in the rush to capitalize on the new technology.  Within 10 years, however, people began to realize that you couldn't place those DNA-based phylogenies in context without some knowledge of basic biology so the field corrected itself, including the new theories and technologies.  I imagine this is what will happen as a result of the current push for hypothesis testing in phylogenetics.


Patrick O'Grady
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Review: Webb et al. (2011) DNA Barcoding of North American Ephemeroptera: Progress and Prospects

10/12/2012

3 Comments

 
Picture
This paper was a convincing argument for the promise of DNA barcoding taking over the world, basically. DNA barcoding of aquatic macroinvertebrates  is gaining backing as an extremely useful tool for taxonomic identification and research, and in turn,  application in bioassessment programs.  Some have argued that DNA barcoding is an unreliable way to identify aquatic macroinvertebrates, but this paper shoots those ideas down; (!!!) as it found that the average intraspecific divergence  was 12.5%, while the average intraspecific divergence was 1.97%. While there were some complications in identification, caused mainly by polyphyly and species complexes (which still need to be further studied and resolved,) in general these results indicate that DNA barcoding is, in general, a promising tool in aquatic macroinvertebrate taxonomy and bioassessment programs. 

Aside from the intra and interspecific  divergences being accurate, for the most part, this paper further points out that DNA barcoding is particularly useful for other reasons.  In addition to helping streamline the identification, delimitation, and discovery of species, DNA barcoding also gives consistent results across life stages, which is particularly important in aquatic ecology applications, as a large majority of benthic macroinvertebrates are immature. In many cases, taxonomy is based on adult male morphology, and identification of immatures, particularly early instars, is exceedingly time-consuming and requires substantial training. Additionally, specimens are often very tiny, and delicate, which can lead, in many cases, to missing gills, caudal filaments or even legs, which can in turn further complicate accurate identifications.  Furthermore, the use of DNA barcoding allows for data standardization, and thus a broader, more accurate  comparison of results.

This paper also suggested that much more work on North American Ephemeroptera  taxonomy and classification is required, as many currently recognized species are  highly divergent. Most of these confused species have complex histories of synonymy and reflect  the 60 year trend in North American mayfly systematics towards inclusive species concepts. Further taxonomic work that synthesizes a variety of identification and classification methods including morphological, biogeographic, ecological, behavioral and molecular techniques is required to test current species hypotheses, particularly of those unusually divergent Ephemeroptera species. DNA barcoding is one of the techniques that will be useful in this aim of achieving stable, supported species hypotheses. Re-examined and updated species hypotheses will allow us to identify aquatic insects more accurately and more efficiently, which will in turn allow us to determine and communicate the ecological characteristics of a species, such as phenology and tolerance to pollutants, and thus  improve our ability to utilize these organisms in bioassessment programs. 

Natalie Stauffer


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Jiji Foundation Grant Funded

1/25/2012

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The lab was just awarded a $3500 grant to do barcoding of aquatic insects in Northern California. Congrats to Brian for all his hard work on this project.
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    Patrick

    Professor
    Cornell University

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