Undergraduate Projects!

Well, that was another two months of frantic work for the Team Aquatic Virus. We’ve been been busy in the lab following up on samples from the January research cruise in the Pacific Northwest. The team’s been joined by senior Victor Biguma, who has been working with fellow student researchers Chaunte Lewis and Mitch Johnson to extract DNA from all sea stars from the cruise, and perform SSaDV qPCR on their tissue extracts to determine load and prevalence. We’re almost there – it does take time!

Elliot’s been busy working with Dan Buckley’s group at Cornell preparing samples for bacterial analysis. He’s taking a closer look at sea star-associated bacteria inhabiting various animal compartments to understand whether their microbiome corresponds with their phylogeny. This is poised to greatly advance our understanding of the disease as well, since we have a huge data set of disease-associated bacterial communities. We also have cultivated 48 strains of bacteria from the sea stars themselves. Unfortunately, no Archaea – repeated attempts have failed to identify any associated with them.

The big news over the last month has been that Ian has engaged his undergraduate class at Cornell (BioMI 3500 – Biological Oceanography and Ocean Biogeochemistry) in sea star disease research. As part of the class, students spend 2/3 of the semester designing and proposing group experiments which they then perform, analyze, write up and present in the last 1/3 of the class. This type of active learning activity is crucial for recruiting budding scientists to the field, and provide students with the opportunity to get their hands wet!

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Starfish arriving to their new home!

The groups proposed three projects to address key, unanswered questions related to SSWD: 1) How does temperature and bacterial load affect SSWD progression?; 2) How does infection with SSaDV influence their microbiome composition?; and 3) How does filtration of aquarium water affect the persistence of SSaDV and progression of SSWD?

After spring break, the students set about performing the experiments. The first challenge: what stars to use? Obtaining animals from the field is logistically very challenging, and it’s not always practical to obtain animals during an epidemic since you don’t know their initial infection status. But, sea stars are easily obtained via the aquarium trade! This provides us with another major advantage: it allows us to test the effects of SSaDV on species that are not currently affected by the disease – and determine if they might be affected should the disease spread into new geographic areas.

In early April, the lab received a shipment of 40 stars from an aquarium supplier, comprising Archaster typicus, Echinaster sp., Protoreaster nodosus and Linckia laevigata. The first challenge in this exercise was to provide them with a “common garden” or “holding tank” – fortunately, the class had received a very large touch tank as a teaching grant a couple of years ago. When the animals arrived, they could be adjusted to room temperature slowly, and the high volume of water meant that levels of dangerous ammonium were kept low.

Test stars in the common garden…

After 24h in the common garden, the students established their treatments. There were a total of 11, 10 gallon tanks representing controls, viral amendments, bacterial challenges, manipulated temperatures, and filtration types. Key variables that students measured included animal health state (using the SSWD 5 category progression), weight of the animals, SSaDV load in tissues and the water column, and finally bacterial community composition, which they tackled using ribosomal intergenic spacer analysis (RISA).

Setting up the gravity filter: (left) establishing the Schmutzdecke and on the right the actual filtration

Experimental setup: Living in Ithaca imposes constraints on the availability of seawater – thanks to Mitch and Elliot, sea stars were kept fresh in artificial sea water!

Weighing sea stars early in the experiment…

And over time, development of SSWD-like signs…

At the conclusion of the experiment, students spent a week dissecting and digesting data. The experiments threw up (as is always the case) some unexpected and interesting results, and it was up to the students to investigate why and how these phenomena occurred. The net results of their experiments were:

  1. Temperature induces SSWD-like disease in Linckia sp, but enrichment with a bacterium native to Evasterias troscheli buffered the effects of temperature (i.e. animals with the bacterium that were subject to elevated temperature didn’t develop SSWD as fast as those with temperature alone;
  2. Infection with SSaDV imparted drastic changes to the overall community structure of the sea star microbiome, and SSaDV appears transmissible to both Archaster typicus and Protoreaster nodosus; and
  3. Gravity filtration and UV treatment reduce overall SSaDV loads in aquarium water, but it is gravity filtration that leads to slower progression of SSWD compared to UV

This afternoon the students presented their final reports to the class, and did a magnificent job. We’ve learned something about SSaDV, SSWD, and the students have learned how to design and execute experiments on the microbial ecology of SSWD in the process. Definitely worth the effort!

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Happy students Samarra Scantlebury, Degan Call and Lu Han after presentations…

Meanwhile, Ian has been busy behind the computer looking at DNA sequences. Like the end of the credits in a Marvel universe film, there is something else in the sea stars that we hadn’t originally noticed. Wait for the next movie to see whether the baddie is really a baddie…

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Riddle me this: What virus has only two ORFs and is over 8000 nucleotides long…


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