Zephyr 2.0 and Mesquite 3.3 released

Yesterday, after about three years in gestation, we released Zephyr 2.0.  Zephyr is a Mesquite package that manages interactions with phylogeny inference packages including RAxML, GARLI, PAUP, and TNT.


The most notable additions to Zephyr 2 include implementation of the SOWH test, CIPRes support, much better interapplication communication, and more extensive support for PAUP.   Many bugs were also fixed, and other improvements made.


The Swofford-Olsen-Waddell-Hillis test allows one to test particular aspects of phylogenetic structure, such as the presence of a hypothesized clade. Given a Mesquite file containing:

  • the data matrix
  • a constraint tree showing just the phylogenetic structure to be tested (e.g., a tree showing just the one clade with everything else as polytomies)
  • the model tree: the best tree with branch lengths that fits that constraint (inferred in a constrained analysis)
  • models of evolution (e.g., GTR+I+G) with parameter values inferred from the matrix,

then the SOWH feature in Mesquite will automatically find the observed value of the test statistic using whatever tree inference program you choose among those Zephyr supports, and simulate data many times on the model tree, calculating the test statistic for each simulated matrix.  It will show you the p-value as the analyses goes along, and gives a report once you have decided you have done enough replicates.


In the figure above, the model tree on which data are simulated is shown in the middle, with the results from the SOWH test on the right.  (In this example, only four replicates were conducted; to get an accurate estimation of the p-value, many more would need to be done.)

More details are in Zephyr’s SOWH test documentation.

CIPRes connectivity

CIPRes (CyberInfrastructure for Phylogenetic Research, http://phylo.org) provides a gateway for doing phylogenetic inference on a fast cluster of computers.  Zephyr 2 allows one to run analyses on CIPRes from within a Mesquite session, and will harvest the results once done and move the trees into the Mesquite file.

Interapplication communication

Zephyr 2 has many improvements under the hood, including much better communication mechanisms between Mesquite and the external program.  Among the improvements are the option to have Mesquite directly start the external program (as opposed to asking the operating system’s shell to do that), which gives Mesquite more control over the process.

Better PAUP support

Zephyr 2 now provides a means to do likelihood, distance, and SVD quartets analyses using PAUP from within Mesquite.

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Happiness is a Big Tree on the Wall


In the hallway outside my lab, about 800 species of Bembidiina, together in one tree.  🙂


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A Stained Trepanedoris

I started learning how to do stained glass pieces over the summer, and for my first piece I decided to honor our Discovering Insect Species class by doing the head of an undescribed species of Bembidion (Trepanedoris) that we discovered.  I finished the piece on Tuesday; here it is:


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Flavor 100

Today we reached a milestone:  our 100th flavor in our approximately weekly Guess the Potato Chip Flavor event in my lab.  Today we decided to celebrate the event not only with our 100th flavor, but also with five other flavors to round out the meal.  Those five other flavors were some current Lay’s flavors:  West Coast Truffle Fries, Southern Biscuits and Gravy, Greektown Gyro, New York Reuben, and Pico de Gallo.  Over the last couple of years we have had quite the variety: Happiness Butter (from Japan, by a South Korean company), Butter Chicken (Lay’s, from Canada), Slow-cooked Ribs (Old Dutch, from Canada), and many others.  Some were excellent, some.. not so.  The two that stayed in the bowl the longest, by far, were Sea Salt Pomegranate and Cappuccino.  We try to stick to potato chips, but we do occasionally branch out into other types (such as corn chips) if there are compelling flavors or if we have no novel potato chip flavors.

But the 100th flavor… now that was a fantastic one.  My graduate student James Pflug acquired it online, from the UK, and, well, here are two pictures:


They have edible gold stars on them!


The flavor:  Winter Berries and Prosecco with Edible Gold Stars and a touch of Fizz, by Marks and Spencer.  Berries and sparkling wine with gold stars.  Wow.  I can’t imagine how we can top that.

Here’s the full list of what we have tried.

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Discovering Insect Species: the Trepaneleven Awards!

In the Discovering Insect Species course, the eleven of us (students Alex, Ana, Danielle, Elle, Julia, Mamo, Shannon, Tom, and Trevin, plus John and me) formed a great team, and we learned a lot together.  Over the course of the term, we became… the Trepaneleven!

TShirt1 small

Design by Julia Amerongen Maddison.

As with any group of superheroes, talents vary from hero to hero.  In order to recognize the amazing abilities and achievements of the Trepaneleven, we held an award ceremony, at which awards were awarded.

And the winners are:

Leaves Analysis Champion: Julia

Julia excelled at finding the best species delimitation using a pre-release version of the Mesquite package Leaves (which attempts to delimit species using gene trees).  Her award was a Leaves Champion trophy.


Most Unusual, Pleasing, and Course-appropriate Career Ambition Award: Elle

Elle’s interest in insect taxonomy and illustration made her the obvious choice for this award.  Her award consisted of a magnifying glass (with built in vial!), some watercolor paper, and some Pigma pens.


Best Dry Sauté and Best Ice Cream Proponent Awards: Danielle

Danielle taught us the wonders of sautéing without oil during our Malheur field trip (as, well, the oil was inadvertently left in Corvallis).  She was also excellent at leading us to ice cream, which the judges very much appreciated.  Her award was a trophy topped with crossing items: a skillet and an ice cream cone.


Most Ambitious Taxonomist Award: Ana

As someone who has described new species in her native Brazil, the country that probably contains more undescribed species than any other, Ana was a natural for this award.  Her project (an interactive key to Bembidion (Trepanedoris)) also swayed the judges. Her award was a copy of E.O. Wilson’s book, The Diversity of Life.


Most Unexpected Comments Award: Alex

Let’s just say you had to be there, in multiple places at multiple times.  With Alex’s interest in forensic entomology, a zombie survival guide and a bug collecting kit seemed the appropriate items to honor his achievement.


Best Habitat Discovery and Best Car Snack Awards: Mamo

Mamo discovered, at Malheur NWR, the habitat of the undescribed species that we first found at Klamath Marsh NWR.  This allowed us to collect what will become the type series of the species.  It was the most significant habitat discovery during the course, and for this the judges will always be thankful.  As important, though, was Mamo’s evangelism for the wonders of Spam sushi, of which we became rather fond on our road trips.  Her award consisted of a book on the history of Spam.


Awards for Solo Discoverer of a New Species, Best Harpalinophile, Best Lover of Words, Best Reaction to a Tick:  Shannon

Shannon not only discovered a new species in her parents’ backyard (it is still the only known locality), she was also overly adept at collecting little harpalines (thinking they might be Bembidion).  She loves words and their histories, and so one way we honored her contributions was with a copy of Brown’s Composition of Scientific Words.  On the Malheur trip, her neck decoration one day, a tick, caused such a reaction that we immortalized that tick in a very small clear box, and presented that to her as well.


Best Knowledge of All Things Aquatic and Best Walkie-Talkie Personality Awards: Tom

Tom knew everything about aquatic invertebrates (at least, that was part of the class myth).  He also was excellent at manning a walkie-talkie so that we could communicate between vehicles as we drove our convoy to and from the field sites; his British accent and humor (or, rather, humour) were a real plus.


Tom was also adept at spreading parts of our aspirators around the countryside, and so his trophy was affixed with one such piece.


Best Effort to Follow Instructions and Not Screw Up Award:  Trevin

Trevin did an excellent job of following instructions.  He also coveted a National Wildlife Refuge sign (containing instructions) that we were given by one of the wildlife refuge managers. In hopes that Trevin would continue to follow instructions, the judges were pleased to give him this award.



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A nematode-filled Bembidion canadianum

Shannon, a student in the Discovering Insect Species course, went down to Eugene last weekend and looked for Trepanedoris. To say that she did well would be an understatement.  She caught all four species that I had seen from the Willamette Valley of Oregon (Bembidion acutifrons, B. elizabethae, B. fortestriatum, and B. siticum), and more.

The real prize was a male of a species I had never seen from this area.  Here’s the live beetle:


At first glance I thought it was an undescribed species, but after looking at it more closely I now think that it is an extremely small specimen of Bembidion canadianum.  I have only ever seen one specimen of Bembidion canadianum from Oregon, collected in 1955 from east of the Cascades, around Upper Klamath Lake.  The specimen is housed in the OSAC.

The specimen Shannon caught was a male with a rather swollen abdomen, and it walked very slowly.  I’ve seen this before, and it usually points to some unpleasantness (at least for the beetle) inside its body cavity, in the form of nematodes.  And, sure enough, when I opened up the abdomen to expose the tissues to ethanol so that the DNA would be well-preserved, there were a lot in there:


Here’s what part of the abdomen looked like; all of the little white tubes here are nematodes:


I’ve seen many nematode-filled Bembidion over the years, but this one was unusual, in that it had nematodes of different sizes: a few big ones, and many small ones.  Normally all the nematodes are of more or less the same size. Another thing that is unusual about this beetle is that it still had what appeared to be fairly healthy testes and accessory glands; a nematode-filled Bembidion normally has very little of its own tissue remaining.

Although interesting, the nematodes will cause some problems in extracting beetle DNA from the specimen – as it will be hard to avoid extracting nematode DNA instead!

Posted in Revising Bembidiina, Z499 (Discovering Insect Species) | Tagged , , | 2 Comments

Discovering Insect Species: preparing beetle DNA

Over the last couple of weeks in our Discovering Insect Species course we have been processing samples from our Klamath Marsh trip, and worked with the DNA of the beetles.  Here’s a bit of what we have done.

Extracting the DNA

We start out with a bit of a beetle in a tube.  This is what it looks like:

A bit of a Bembidion in a tube.  This doesn't happen to be a Trepanedoris, but it would look just about the same if it were.

A bit of a Bembidion in a tube. This doesn’t happen to be a Trepanedoris, but it would look just about the same if it were.

We then went through all of the steps required to dissolve the soft tissue in that sample, and from the soup that results extract the DNA by itself into a clean tube.  Here are pictures of a couple of those steps:

JuliaDNAExtraction ElleDNAExtraction

In the end, we get beetle DNA in water (plus some salts).  It looks just like normal water, but there is beetle DNA in there:


The students extracted DNA from a total of 36 specimens, most of which they had collected on our Klamath Marsh field trip.

Amplifying a gene we want to study

Once all of the DNA in the beetle was extracted, we then wanted to pick a few genes, and make many copies of those genes so that they could be sequenced.  The first gene we picked was 28S.

Everyone combined various chemicals in some small tubes.  These chemicals included nucleotides and an enzyme, Taq polymerase, which copies DNA.  We also added short stretches of DNA called primers that match part of the sequences for the 28S gene in the beetles, and that will providing a starting point for the reaction.  The end result will be that the enzyme will produce many copies of the beetle’s DNA between the two primer regions on the chromosomes.  This reaction is called the Polymerase Chain Reaction, or PCR, and is a standard way of making many copies of (“amplifying”) DNA.

Here we are getting small tubes set up for PCR:

preparing PCR

The tubes are then put onto a machine called a thermal cycler, which once closed and started will go through various cycles of changing temperatures:


In the end you get, for each sample, a little tube that contains zillions of copies of the piece of the 28S gene between the primers:

PCR product

At least that’s what we would hope would happen.  To confirm that the DNA piece was amplified for each sample, John then mixed a small amount of the liquid from each PCR tube with a dye that will attach to DNA and fluoresce under UV light, and ran the combination on a gel, thus allowing us to see whether DNA was amplified:


Below is a part of the gel showing the 28S PCR products from the amplification done by Tom and Shannon (on the left) and Mamo and Julia (on the right):


The first eight bands are all good, and indicate that we successfully amplified the 28S gene for those beetles.  The ninth spot is dark as that is the negative control – we did everything for that PCR as we did for other PCRs, but no DNA was purposely added.  If there had been a band there, it would suggest some DNA accidentally got into the chemicals, which would be bad.  The next eight spots are the same, but for a different set of beetles, and the last spot is another negative control for that second reaction.

Here’s John explaining that the 28S reactions were quite successful!


We also did PCRs for the COI gene during the same sessions.  Two genes (28S and COI) for 36 beetles combined mean that the students will have produced a total of 72 DNA sequences.  The next step is getting the PCR products ready to be sequenced, which I will report on in a future blog post…

Posted in Revising Bembidiina, Uncategorized, Z499 (Discovering Insect Species) | Tagged , , , | 2 Comments