Two ŔÖ˛Ą´«Ă˝ College scientists have received nearly $373,000 from the National Institutes of Health for first-of-its-kind research into the genetic functioning of bacteria that cause Lyme disease.

The project draws on new capabilities in high-resolution microscopy that make it possible to observe spatial arrangements of RNA in Borrelia burgdorferi, one of four Borrelia species that cause Lyme disease. This study of variations in RNA location and shape could eventually suggest new medical responses to Lyme, which strikes up to 300,000 people in the U.S. annually.

Receiving the $372,639 grant for the two-year project are physicist Travis Gould, an expert in the field of fluorescence nanoscopy, and biochemist Paula Schlax, who researches gene expression in spiral-shaped bacteria, also known as spirochetes, such as Lyme bacteria.

Biochemist Paula Schlax and physicist Travis Gould pose with Gould’s STED super-resolution microscope in Carnegie Science Hall. (Phyllis Graber Jensen/ŔÖ˛Ą´«Ă˝ College)

“RNA is an intermediate in the process of cells making proteins,” says Schlax, a professor of chemistry and biochemistry at ŔÖ˛Ą´«Ă˝. “We’re trying to understand generally how production of proteins gets turned on and off when the bacteria move from ticks to mammals and from mammals back to ticks” — changes in the bacteria’s environment that change the shape and location of RNA.

“We know from other bacteria that RNA’s location inside the cell seems to affect how long that RNA lasts — whether it’s near the edges of the cell, or the ends of the cell, or spread out evenly inside. Our hypothesis is that how fast RNA gets broken down, or doesn’t get broken down, probably helps the cell decide which proteins to make when conditions change, such as when the bacteria moves from the tick to a mammal or vice versa.”

Variations in protein production could cause variations in the bacteria’s disease-causing capability. “The more we understand that process,” Schlax says, “the easier it is to think about new targets for drugs and new therapeutics.”

Until quite recently, the physical limitations of microscope technology curtailed its usefulness in testing such a hypothesis. The bacteria have a characteristic shape, says Gould: very skinny in relation to length. The length is typically around 20 microns, or millionths of a meter, but the bacteria’s internal diameter is vastly smaller, at about 200 nanometers, or billionths of a meter. (A piece of paper is about 100,000 nanometers thick.)

“That 200 nanometers is, in the best-case scenario, at the limit of a conventional microscope’s resolution,” Gould says. “So a conventional microscope can’t answer these questions about where RNA is within that 200-nanometer cylinder.”

But Gould, an associate professor of physics, is an innovator in imaging technologies that use lasers, fluorescing molecules, and other means to attain much higher resolution. For the NIH-funded research, he has adapted an existing ŔÖ˛Ą´«Ă˝ microscope that he built and that uses a process called stimulated emission depletion, or STED, to capture images of the B. burgdorferi RNA.

Going from older microscopy technologies to Gould’s newly updated STED “is like putting on glasses for the first time.”

Specifically, he added another laser to the instrument that expands its imaging capability from two to three dimensions. Going from older technologies to this latest iteration, says Schlax, “is like putting on glasses for the first time.”

Complementary to the STED technology, the researchers and their students will use a technique called fluorescence in situ hybridization (FISH) that deploys fluorescent probes to specific parts of the “transcripts” that the microbe’s DNA imparts to its RNA.

The research will be the first to identify patterns of transcript localization within B. burgdorferi, and, notably, the first research to use STED microscopy for this sort of localization within any spirochete.

This is significant given the range and impact of diseases caused by such bacteria, including syphilis, yaws, periodontal disease, and leptospirosis, whose effects include kidney failure.

Joining Schlax and Gould in the project are ŔÖ˛Ą´«Ă˝ students and research associate Anna Bowsher, whose position is funded by the NIH grant. The work entails growing B. burgdorferi microbes in the lab, affixing individual cells to slides, and introducing DNA molecules, complete with fluorescent tags, that are tailored to activate a specific RNA response.

Then the slides will be examined with Gould’s STED microscope, and the results compiled into a spatial-distribution analysis of different types of RNAs. The team hopes that they will have results to report by summer 2020.

The project will involve both thesis students advised by Gould and Schlax and students doing summer research. “These kinds of projects really are great for students to see how science is done,” says Schlax, “and hopefully get their names on some papers and keep them interested in science.”

With the use of fluorescing molecules now standard practice in high-resolution microscopy, STED imaging achieves enhanced resolution through a technique of selectively switching off such molecules. STED is one of a number of so-called super-resolution techniques developed to bypass the diffraction limit, a limit on the resolution of conventional microscopy imposed by the length of light waves.

Gould estimates that all told, there are likely two dozen or so labs equipped with commercially available STED microscopes, and another handful that use custom-built instruments like his.

Caroline Holm ’17, Southern Maine Community College intern Alex Krech, and Anna Berenson ’16, work with the bacteria that causes Lyme disease in the lab of chemistry professor Paula Schlax this past August. (Phyllis Graber Jensen/ŔÖ˛Ą´«Ă˝ College)

Lyme disease is just awful. In addition to being transmitted by disgusting blood sucking ticks, the disease signals its arrival with spreading, bullseye rashes.

It often culminates in joint pain, headaches and other debilitating symptoms, and upwards of one person in five develops long-term symptoms, including chronic pain, vision problems and heart complications.

The three-year, $345,750 grant from the National Institutes of Health will also support research opportunities for ŔÖ˛Ą´«Ă˝ students.

But from a scientific perspective, Lyme disease is fascinating, as ŔÖ˛Ą´«Ă˝ chemistry professor Paula Schlax explains. The bacteria that causes it, Borrelia burgdorferi, is unlike any other organism she’s ever studied. “It has features and characteristics that are unlike most other bacteria,” Schlax says.

“It doesn’t require iron to live, and it may be the only organism for which that’s true,” she adds. “Most bacteria have circular chromosomes, a big loop of DNA, but this has DNA that’s linear. We have very little information about how genes get turned on and turned off in this organism.”

Paula Schlax is a professor of chemistry.

Schlax’s study of Borrelia, its genes and the “switches” that control them, has earned her a three-year, $345,750 grant from the National Institutes of Health — funding that will also support research opportunities for ŔÖ˛Ą´«Ă˝ students.

The goal is to better understand how these switches work when the bacteria moves from a tick into a mammal, and from a mammal into a tick.

Although antibiotics often succeed in treating the disease, more than half of Maine’s counties are at high risk for Lyme disease, which is spread by deer ticks.

In Maine, 2014 was a record year for Lyme disease, with nearly 1,200 confirmed cases. The six New England states accounted for 35 percent of the nation’s confirmed cases of Lyme disease.

Every year, roughly 30,000 Americans are confirmed as having been infected with the Borrelia bacteria. But because Lyme disease is often not reported or diagnosed, the actual number of infections could be 10 times higher than that, according to the Centers for Disease Control.

“I had a couple friends in college whose mothers got Lyme disease and they both had very, very serious complications from it,” Schlax said. “So it’s always been in the back of my head that this can be really horrible for people.”

Schlax started working with Borrelia when she received funding through Maine INBRE (IDeA Network of Biomedical Research Excellence) in 2009. The Maine INBRE award allowed her to focus on understanding gene regulation in Borrelia.

The infection that causes Lyme disease is activated in the bacteria when it moves from tick to mammal. Understanding how those switches occur — or better yet, figuring out how to stop or alter them — could yield new treatments for Lyme disease, Schlax says.

“The more we understand that, the more likely it is that someone else can use that information when thinking about ways to alter that transmission to make [the bacteria] less effective,” she says.

Schlax’s research examines how the shape of an RNA molecule influences its ability to interact with other molecules in the bacterial cell. In her lab in Dana Chemistry Hall, she and her students will use the grant to monitor how fast Borrelia RNA gets broken down by enzymes called ribonucleases, important information to know when studying how to inhibit the bacteria.

“We’re interested in interactions that are responsible for enzymes that chew up the RNA into little nucleotide pieces,” Schlax says. “How does that work?”

“It’s always been in the back of my head that (Lyme disease) can be really horrible for people.”

Using living cells, Schlax and her students add antibiotics to stop Borrelia from making new RNA. Then, at various intervals, they measure how much RNA has been broken down by the ribonucleases in host cells.

“Sometimes how tight [the proteins] bind is what matters, and other times it’s who binds first,” Schlax says. “I’m interested in switches that work by both of those mechanisms.”

The NIH grant will allow Schlax to hire a lab technician as well as fund paid summer research opportunities for as many as nine students over the life of the grant. In addition, three thesis students are currently working with Schlax on related research.

Students “will be measuring the protein levels, RNAs, they’ll be making synthetic pieces of RNA to see what the ideal piece of RNA to be cut is,” Schlax says. “They’ll be doing everything that’s in the grant. It’s truly collaborative with them.”

Schlax has taught at ŔÖ˛Ą´«Ă˝ since 1998, and was promoted to full professor in August. She graduated from Clarkson University in 1989, and obtained a doctorate from the University of Wisconsin in 1994.

A ŔÖ˛Ą´«Ă˝ College junior from Maine and a senior from New York state presented biochemistry research to a U.S. representative from Maine this month as part of “Posters on the Hill,” an annual event designed to remind Congress of the importance of undergraduate research.

Both majoring in biological chemistry, Joshua Linscott of Portland and Nicholas Swerdlow of Hastings-on-Hudson, N.Y., were among 78 undergraduates from across the nation to take part in the May 4-5 event sponsored by the , a national organization that supports student-faculty collaborative research and scholarship.

The ŔÖ˛Ą´«Ă˝ pair presented genetic research involving the bacterium that causes Lyme disease, Borrelia burgdorferi. Working with Paula Schlax, associate professor of chemistry at ŔÖ˛Ą´«Ă˝, they studied messenger RNA that regulates the expression of a protein connected with the microbe’s transmission from ticks to mammals.

“The genome of this bacteria is quite unusual,” says Schlax, “and understanding basic principles of protein synthesis will be essential in understanding its life cycle and its ability to be an effective pathogen.”

The gatherings give undergraduate students and their professors a forum to explain their research to members of Congress and their staffs, and to discuss the importance of such research and federal financial support for it.

Linscott and Swerdlow met Maine Rep. Michael Michaud, who represents Maine’s 2nd District, which includes Lewiston. “The Congressman has an excellent relationship with ŔÖ˛Ą´«Ă˝ and was very receptive to our visit,” Swerdlow says. “He asked about our research and looked at our poster, but the meeting was focused on undergraduate research in general.

The pair emphasized that grants from the Maine IDeA [sic] Network of Biomedical Research Excellence, funded by the National Institutes of Health, have supported their research at ŔÖ˛Ą´«Ă˝, Linscott says. “I think my voice represented the thoughts and needs of many undergrads. Undergraduate research is hugely important in preparing students to enter grad school or the job market.”

Described by Schlax as a hard-working, fastidious student, Swerdlow wants to pursue a career combining research and medical practice. He has spent the last two summers performing clinical research on obesity.

Linscott has worked at IDEXX, the Westbrook-based veterinary diagnostics manufacturer, and hopes to do pharmaceuticals research after ŔÖ˛Ą´«Ă˝. Schlax described him as “organized, careful and motivated.”

For all of the students, Posters on the Hill included opportunities to network, see some sights in the nation’s capital and share their research.

“It was amazing to be surrounded by reminders of the history of our country,” Linscott says. “For all of us, I think, it was very freeing to be able to explain our research in depth to other students who could really understand what we were doing.”

Using traps baited with peanut butter, a ŔÖ˛Ą´«Ă˝ research team spent the summer catching small mammals and studying the ticks they carried, specifically looking for ticks infected with the Lyme bacterium. Working with visiting biology professor Ronald Barry were biology majors Elizabeth Rogers ’09 (shown above picking the critters off a white-footed mouse) of Mansfield, Mass., and Nelish Pradhan ’10 of Kathmandu, Nepal. The research was supported by the Howard Hughes Medical Institute. [More…]