In MB&B 320, rewriting the tree of life

It’s been two years since molecular biophysics and biochemistry professor Scott Strobel’s famed brainchild — MB&B 320: “Rainforest Expedition and Laboratory” — was born. Two treks through the Amazon River basin and hours of lab work later, its students have more than just spring-break snapshots of the rainforest to show for it.

The group of undergraduates who took his course has discovered dozens of previously uncharacterized microorganisms — many of which diverge genetically from any known species — that may have useful therapeutic applications for a whole host of human diseases.

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Among the some 500 Amazonian organisms called endophytes — fungal and bacterial microorganisms that inhabit the inner tissues of plants — found and cultured back at Yale, “almost every student found something wildly off the charts,” Strobel said.

Jina Chung ’10 and Daniel Marks ’09 found fungal species that produce compounds appearing to selectively kill HIV and cancerous cells, respectively.

Three of the endophytes Kathleen Dantzler ’10 has spent the last months characterizing inhibit trypanosome, a protozoic parasite that causes African Sleeping Sickness.

Cong “Carl” Ma ’08 discovered a third fungal endophyte with anti-oxidant properties.

Meanwhile, Sun Jin Lee’s ’09 organism of interest inhibits inflammation in human placental tissue and prevents apoptosis, or programmed cell suicide.

Ten of the new organisms the undergraduates found were so genetically divergent from known species of endophytes that they became the first members of new taxonomical genera, Strobel said. He explained that these 10 were between 15 and 30 percent different from any sequence found in GenBank, an online database that compiles the genetic sequences of known organisms.

Lee admitted to the thrill of baptizing the genus and species names of her “own” organism. She finally settled on “Aurosphaeria radiaflava” — which, translated, means “golden spheres … radiating yellow” — in honor of the microbe’s physical characteristics.

The students also constructed phylogenetic trees to measure each of the microorganisms’ phylogenetic distance from its closest known genetic relatives. Assays conducted on each cultured endophyte to record bioactivity revealed that 65 were bioactive.

The students’ findings, which were released the August 25 issue of PloS ONE, speak of the untapped scientific potential of places like the Amazon, which has the highest density of biodiversity in the world, Strobel said. Currently, only one-fifth of the projected 200,000 species of existing fungi have been discovered, and research on endophytes remains a fairly undeveloped field.

“The fact that 18 untrained undergrads were able to discover whole new collections of fungal species and genera speaks to the need for more investigation in tropical areas,” he said. “The incredible diversity [in the Amazon] is such that it’s almost trivial to discover a new species.”

Discovering Diversity

Students collected their specimens in March and have spent the intervening months isolating and culturing the organisms, sequencing their DNA and assaying them for biological activity.

This final step — the final assignment for the course — is possibly the most important, Strobel said.

Strobel required course enrollees were asked to find a science faculty member with a biological assay that could be screened against the endophyte-extract library the students created over the summer. They then each assayed their compound of interest — one that they had isolated from the library based on their research interests.

“But unlike the typical situation where a student knocks on a professor’s door hoping to be assigned a project, in this case the student comes up with a unique resource that could significantly advance the program of the faculty member,” Strobel said.

Lee remembers the experience as “really scary.”

“These assays are fishing in the ocean. You don’t know if you’re going to have a hit,” she said. “So you’re asking all these professors to spend their time, money and resources on this random fishing expedition.”

Before their excursion to the rainforest, Strobel had asked his students to prepare a plant-collecting list drawing upon on a defined theme. Lee said hers had been women’s health issues, based on which she collected plants with medicinal properties relating to diseases and conditions that affect women.

“But half of them were completely random — things I thought looked interesting,” she said.

But get a hit she did.

At a lab in the Obstetrics and Gynecology department of the School of Medicine, Lee found a professor willing to do an assay with her 46 samples. The lab works with discarded placentas from C-sections performed at Yale-New Haven Hospital, studying compounds that incite inflammation in the placental-fetal membrane. Their research is crucial to understanding pre-term labor — caused by inflammation due to infection in 30 percent of cases — which can lead to the delivery of premature babies, Lee said.

Lee’s assay screened for a compound released by the endophyte samples that would suppress this over-reactive inflammatory response system. Lee found a single such hit, one that came from a carnivorous plant — not, as she had predicted, from one of the plants she had collected for their medicinal properties.

Dantzler, who assayed her samples at a parasitology lab at the medical school and also found a hit, said she had not expected to find a compound with so much therapeutic promise.

She recalled her surprise the moment when she first introduced her culture into a sample of parasites to find that it entirely killed them.

“Usually, the media has reddish color and the parasites are oval-shaped, but the media was this orange, cloudy color and looking under the microscope, the parasites had all become sickle shaped,” Dantzler said. “My post-doc was so excited. He was running around the lab and showing it to everyone.”

A love for research

While discovering a new species may seem like the finale of a scientific journey, Strobel emphasized that the most difficult part still lies ahead for MB&B320 students.

Now that they have discovered that their organisms of interest may exhibit potentially useful biological activity, they must now use techniques in organic chemistry, such as biological fractionation and X-ray crystallography, to purify and characterize the chemical compound responsible for that activity.

Lee’s on her way to putting the finishing touches on exactly that. Having separated the components of the sample using column chromatography, she has determined the crystal structure of her bioactive compound and identified it as the known substance Breselzin A, which stops protein trafficking from the endoplasmic reticulum to the Golgi apparatus. Lee said she anticipates her findings will be published sometime next year.

Blair Benham-Pyle ’10 is at a slightly earlier stage in her project: She has partially isolated a compound effective against the bacteria responsible for Staph infections, a common infection among immunosuppressed patients.

Now the challenge remains to obtain crystal structure for the compound through further purification.

Meanwhile, Dantzler’s immediate goals are to test her compound in human cells and against the malaria parasite, which, like African Sleeping Sickness, is caused by a protozoa.

But the course — both Strobel and the students interviewed stressed — has been about more than just the end result.

The original idea, Strobel said, was to give students a chance to think and act like scientists: to depart from the “recipe-style” format of many lab courses, by giving them the chance to make their own discoveries.

“We want to give students the sense that they can still contribute without being world experts — and making those discoveries is not so daunting,” he said.

Dantzler— who said she “knew [she] had to take” Rainforest Expedition and Laboratory by the second day of shopping week — said the course offers the unique ability “to follow a project from beginning to end.”

Students are self-directed from the get-go: they identify their interests, collect their own plants and then characterize their own microorganisms, she explained.

“It feels like it’s ours,” she said, of the research.

Lee, on the other hand, said she took the class on a whim, and committed “completely last-minute.”

Having previously worked at a research lab at Yale, she said she was beginning to be turned off by the disconnect between research and the real world.

“I was finding more and more that I felt like ‘science is all done’ and ‘research isn’t exciting,’” she said. “I needed something more relevant to people’s lives.”

Through the course, Lee rediscovered her love for research — especially clinical research, which appealed to her because of its faster turnover rate and greater degree of interaction with patients.

“It ended up redeeming my faith in science,” she said.

Pyle said she in large part chose Yale over other colleges precisely because of “Rainforest Expedition and Lab” and her correspondence with Strobel before her formal acceptance.

“It’s honestly really lived up to my expectations,” she said of the course, adding that she has decided to become an MB&B major since coming to Yale. “I found a new genus. I found a new species. I’ve found a new drug that could potentially cure lives.”

She added with a chuckle, “I have enough research to keep me occupied for several years.”

Comments

  • Alum

    Wow, this is an awesome course. I hope Strobel and MB&B can keep it going!