"What would the world be, once bereft Inconceivably, unerringly, indisputably, I have arrived at the end of this REU program. Our submissions of specimen records, DNA sequences, chromatogram trace files from the raw output of the sequencing process, and identifications have been uploaded to the Barcode of Life Database. My poster is printed, and as you read this, I have presented at the OIMB REU symposium and already flown home to North Carolina. This has been a deliriously wonderful summer, replete with late nights pipetting precise, minute aliquots of strange, clear liquids into other, precisely measured clear liquids. I have learned every step of the genetic sequencing process, from extraction, to PCR, to PCR, to re-PCR, to troubleshooting PCR, to one final PCR with fresh primers that works like a charm, to agarose gels, to loading plates, to cleaning and processing sequences, to making the data accessible on a public database. I found out how easily mistakes creep in, and how to root them out with meticulous records and by double-checking every step of data management and analysis. I have discovered an entire phylum which I had barely heard of before this internship, and though hardly an expert, I can catch and identify a couple of genera, and I have learned the basic taxonomic divisions of Nemertea. I have also submitted an abstract to a conference for the first time, and I am planning to present my research at the Society of Integrative and Comparative Biology's 2024 meeting in Seattle! More to the point, by my nine weeks of effort, I have documented 44 species from the Red Sea, and demonstrated that there are over a hundred more that are waiting to be discovered. Never before have I been so close to the cusp of new discoveries, nor more acutely aware that there are uncharted universes of biodiversity, in all its wriggling, crawling, squirming, sessile, chemosynthetic, carnivorous, parasitic vivacity, propelled by the simple fact of natural selection to unimaginable variety. There is so much that is known, and yet so much more lies just over the horizon, or perhaps just under the water's surface. Of course, this summer would not have been one fifth the joy that it was without the support and camaraderie of my fellow REUs. Their positive qualities are too numerous to recount, but rest assured they are each unique and wonderful, and have excellent blogs that you can peruse! I am beyond grateful for the opportunity to meet all of them and explore this pocket of Oregon together. Be happy and healthy. "Something told him that something was coming to an end. Not the world, exactly. Just the summer. There would be other summers, but there would never be one like this. Ever again.
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"I'm wandering in the mist just now. It'll blow off in a bit." -Margaret Renkl, Late Migrations Last weekend was an event I had been looking forward to, our public presentation at the Charleston Marine Life Center. All the REUs who are still here (Madison, Randi, and Tara are off having cool adventures) tabled for a couple of hours to talk to visitors about our research projects. Audrey and I assembled a tableau of live, locally collected nemerteans, an unintentionally devious image sorting game, and a blackberry DNA extraction station. I enjoyed trying to communicate what, precisely, I have spent the last two months doing, and I hope the kids we met enjoyed seeing the worms (and in a couple cases, touching them). The DNA extraction and micropipette seemed to be a big hit, although I do wonder how well we explained the purpose of the exercise. Alas, the rest of my week was not so enjoyable. I came down with COVID on Sunday and spent the next few days moping about and generally feeling crummy. As I write this on Thursday, my fever is gone and I'm feeling much better, but I didn't have a very productive week. Because most of our remaining work is database submissions and polishing posters, I was able to do some work remotely, and Audrey kindly did all the sequence clean up on the lab computer for our final plate. Despite the setbacks, I joined in the poster review and critique session remotely on Wednesday, and I got some helpful thoughts from Dr. Watts, Dr. Emlet, Riley, and my peers. Other than that, I've just been fiddling with R code to compile all of the specimen data and create nice-looking graphs for my poster. Besides my poster, the one silver lining to this week was that I finally took the time to figure out how ggplot controls all the visuals and aesthetics of graphs, and I am excited for future opportunities to develop illustrative plots. It's a little difficult to believe, but I am approaching my last week at OIMB this summer. Our final weekend will be spent touching up our posters, and then Audrey and I simply have to clean up the lab, organize our data sheets in a decipherable system for future reference, give a nice poster talk, and they'll kick us out! I've never had such an intense, focused, and joyful summer, not to mention the gorgeous, sprawling land and sea that surround me, and I already know that I will miss this place, a restful eddy before the current carries me to new challenges and adventures. I love it, for one week more. "I do not know what I may appear to the world, but to myself I seem to have been only like a boy playing on the seashore, and diverting myself in now and then finding a smoother pebble or a prettier shell than ordinary, whilst the great ocean of truth lay all undiscovered before me." -Isaac Newton This week marks the end of our lab work this summer. We re-amplified samples which previously gave us low quality reads (which was, in a pleasant surprise, quite successful), and measured the concentration with fresh Qubit fluorometer standards. This machine is fairly straightforward to use -- DNA absorbs light at a known frequency, and by measuring how much light is absorbed, the fluorometer can measure even very low concentrations of nucleic acids or proteins (our assay tubes ranged between 0.0100 and 0.7000 ng/μL). With these precise measurements, Audrey and I were able to dilute each sample accordingly, load our final 96-well plate, and send it to the OSU Core Lab for sequencing. We must tie up our project, perhaps sooner than we would have chosen, but we have hammered an enormous dent of knowledge into the previously pristine ignorance of nemertean diversity around the Arabian Peninsula, to the tune of 33 novel OTUs and counting. In order to make this information accessible and useful for future researchers, collectors, and naturalists, we have been uploading our data to BOLDSystems, a database meant for DNA-barcodes from all life on earth. One interesting aspect is that BOLDSystems has its own algorithm to delimit OTUs, which is less conservative than our ASAP analysis, so it records a few extra species. This is an important reminder that DNA-barcoding of the COI gene is a good proxy for differentiating species, but it is not the determining factor. The biological species concept is based on whether individuals and populations can produce viable offspring, and populations which do not interbreed will diverge genetically, but there are few sharp lines in the sprawling, patterned chaos of biology. Also outside of the lab, Audrey and I have been trying for the past week to collect nemerteans four our upcoming presentation at the Charleston Marine Life Center. We went to the Barview mud flats with Randi, but we had little luck. After an hour of sieving, we found one medium Cerebratulus, which we broke, and a few tiny, indeterminate worms. Most ended up being annelids (the bane of my nemertean-hunting existence!), though one piece turned out to be a broken-off nemertean midriff, probably from Maculaura alaskensis. It was fascinating under the microscope, with its branching brown intestines, and squeezed in between each branch are ovaries, bulging with eggs, each dotted with a nucleus. The Cerebratulus broke quite catastrophically, and its proboscis squirmed out the gaping hole in its body, but nemerteans have extraordinary regenerative abilities, and we hoped it would survive. This was not to be, for we found it rotting a few days later and dumped it down the sea table drain. A trip the next day to Sunset Bay brought us the classic Tubulanus ruber, but little else. On Dr. Maslakova's advice, we went once more to the mud flats, seeking small, brown, filamentous nemerteans that slink back to their holes as the tide recedes. We found one... and it broke and soon died. I now understand all the complaints I read from the 1800s about worms fragmenting before they could be properly studied. However, we went out on the search one more time today, this time with Dr. Maslakova. We chiseled barnacles off the dock pylons, and discovered quite a few Emplectonema viride. We are very pleased with these specimens, and will bring them to our presentation. I am also grateful for the opportunity to learn about collecting nemerteans, because there aren't many other people in the world who could show me so much about the practice. This week was also the annual Invertebrate Ball, which was a blast! We dressed up as cool invertebrates, and there was a swarm of impressive costumes. I am the proud winner of the Best Invertebrate Walk/Crawl/Strut/Move award. With lab work finished, these last two weeks will be a whirlwind of producing a poster, identification guide, and database submissions, as well as cleaning up our data, spreadsheets, and DNA samples to leave everything in neat order for future documentation. I'm both nervous and excited! Certain Knowledge from Herodotus These are the facts about the fish in the Nile: My favorite part of the past week was interacting with the South Slough National Estuarine Research Reserve. I went with Naia, Audrey, and Taylor to run an informational booth at the Coos Bay Farmers Market. We discussed the invasive five-spine crab (green crab), as well as the reserve's various programs and restoration work. I also enjoyed exploring the market, where I bought some nice plums and met Side of the Tide, the South Coast Folk Society's Morris dancing troupe, who seemed to be having a great time. Thursday morning, I woke up early to help South Slough collect eelgrass seeds. We tromped over to the Barview mudflat, which is just across the slough from OIMB. It is not heavily trafficked, but it was a beautiful and productive site. My duty was to extract mud cores along two 100 m transects and plop them into Ziplocs, and then sieve out the organic material from each one. I was very grateful for the borrowed wetsuit which kept me clean and (mostly) dry. I also saw several of the Morris dancers kayaking past, which was a pleasant surprise.
On Wednesday, we received the sequences from the two plates that gave us so much trouble last week. Our rate of high quality sequences was lower than before, but three-quarters were still good enough to process, which hopefully foretells more species discoveries. We received new standards for our Qubit fluorometer, although we haven't tested them yet, which we hope will allow us to measure the concentration in samples which did not sequence well.
We have also been submitting sequences and associated specimen information to BOLDSystems, but have encountered a few processing issues. The biggest contention is that they would like to simplify our taxonomy, by moving information about which operational taxonomic unit a specimen sorts to or which species it resembles to the notes column, instead of the species column. We'd rather keep this information prominent, because it is important to have a unique identifier for every operational taxonomic unit. The open taxonomy notation ("this specimen is similar to such-and-such species, that one looks like that species") is necessary, since so many nemerteans are undescribed, and we have no other way to anchor unique identities. As we head towards Week 7 of 9, I am feeling the time crunch -- I want to collect all possible data from every sample, submit information to databases, create detailed ID guides, and prepare engaging presentations for the general public and for our poster session, not to mention explore the wonderful Oregon coast, but our time here was always going to be limited. I hope I will find a satisfying conclusion to my project and appreciate the last few weeks for all they are worth. |
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