July, 2021 — Be it bird, bug, or bacterium—no organism on Earth exists in isolation. We all live and interact with other living things, and these relationships between species shape the world around us.
This natural harmony is easy to picture on a global scale: exotic creatures eating fruit in a lush and colorful rainforest, tall cacti thriving under the brutal desert sun, or bumblebees spreading pollen in a field of wildflowers.
But what about the many “invisible” ecosystems that make you you?
Like Earth, the human body is host to an eclectic array of communities, each containing many interacting microbes. These tiny ecosystems, called microbiomes, influence our physiology in myriad ways. In particular, the interplay between the microbiota in the human gut—many trillions of bacteria, viruses, and fungi—plays an outsized role in shaping our health.
Despite its importance, researchers lack a mechanistic understanding of how the gut microbiome as a whole is greater than the sum of its parts. In other words, to what extent do individual microbes influence us, and to what degree are these impacts determined by the interconnected and overlapping interactions between unique species? Studying the underlying biology that governs these relationships is absolutely critical to advancing our understanding of human health. However, the sheer number of different microbes in our gut presents a challenge to cataloging and understanding the effects of their synergy.
Using the natural simplicity of the fruit fly gut microbiome as a model system, the Ludington Lab at the Carnegie Institution for Science’s Department of Embryology is starting to untangle this complex web of interactions.
Electron microscopy of a cross section of the fruit fly gut. The bacteria are seen as dark, round shapes. They are separated from the fly cells by a light gray band of an unknown extracellular substance. // Ren Dodge & Mike Sepanski.
Live imaging of a fruit fly gut. The Bellymount System, developed by a collaboration between Dr. Lucy O’Brien and Dr. KC Huang of Stanford University and Dr. William Ludington of Carnegie, allows researchers to peer into the live tissue of the fruit fly gut in real-time. // Courtesy of Leslie Koyama and Lucy O’Brien.
Handling live flies is a delicate task. Lab manager Daniel Martinez uses a CO2 pad and custom “grabbing” device to prepare fruit flies for an experiment under sterile conditions.
Johns Hopkins University rotation student Gemechu Mekonnen prepares flies for the “Smurf assay.”
Above, rotation student Gemechu Mekonnenon collects fruit flies to analyze their gut permeability. Called the “Smurf assay,” he will feed the flies blue-dyed food. If the entire fly turns blue to the naked eye, Mekonnenon knows bacteria have weakened the integrity of its gut.
“With other kinds of microbiome experiments, the researchers are asking ‘what’s there?’ But we know exactly what’s there. We’re saying, ‘we have this specific system, and when we put this in or take that away, we can see the effect come and go.”
Drosophila melanogaster (the fruit fly) shares 60 percent of its DNA with humans. // Sanjay Acharya
The spark that led to Ludington’s innovative work came shortly after joining the Department of Embryology, where he began to collaborate with Director Emeritus (and current Staff Scientist) Dr. Allan Spradling—one of the founders of modern fly genetics. In the 1980s, Spradling pioneered research and techniques that led to a much greater understanding of fruit fly biology, including manipulating its genome. In no small part because of Spradling, the fruit fly is now the most ubiquitous and powerful model organism used to understand human biology; directly responsible for six Nobel Prizes in Physiology and Medicine.
Dr. Allan Spradling at work in his lab at the Department of Embryology. Circa 1980.
Since starting operations in mid-2018, the Ludington Lab has produced results at an explosive rate, honing in on individual gut bacteria interactions and their effects on targeted biological processes—like the immune response.
Each vial in the Ludington Lab’s incubation chambers holds a different genome of fruit fly.
In the image above, Ludington reaches for a vial from one of his incubation chambers. Each vial contains a different genetic stock of fruit fly—some are wild-type, with no known mutations, and others have lab-induced mutations in a specific immune gene. By comparing the distinct effects of the same bacterium in both types of fly, Ludington and his team can study how it affects the immune system.
But it’s not enough to only understand and manipulate the genetics of the fruit fly. To have complete control over the equation, the lab must also turn “on” and “off” specific genes and proteins in the bacteria themselves—and know where to find them.
Below, postdoctoral fellow Karina Gutierrez Garcia uses advanced bioinformatics to analyze the genetic and evolutionary history of Lactobacillus plantarum—the most common bacteria found in wild fruit flies.
Karina Gutierrez-Garcia studies the genetics of Lactobacillus plantarum. Each purple and gray line to her left represents a separate genome of the bacteria. Her right screen displays a graphical analysis of its evolutionary tree.
“We are working to characterize the fly genetics of colonization—and also the bacterial genetics of colonization.” – William Ludington
Predoctoral associate Kevin Aumiller uses a custom fluorescent lightbox to count bacterial colonies from a single fly. Measuring the number of bacteria in the host is one of the most common aspects of the lab’s experiments.
In the first of the two images below, postdoctoral associate Robert Scheffler displays colonies of fluorescent-labeled Lactobacillus plantarum. By expressing the mCherry protein, the lab can turn the bacteria a pinkish-red color—allowing for easy visualization under a microscope. In the second image, Johns Hopkins Ph.D. student Haolong Zhu analyzes advanced microscopy data from a fly gut colonized with mCherry-expressed Lactobacillus.
By tagging bacteria with a fluorescent protein, researchers in the Ludington Lab can easily locate it in the gut. Top: Robert Scheffler. Bottom: Haolong Zhu.
Below, senior research associate Ren Dodge dissects a fly to prepare a cross-section for imaging at Embryology’s microscopy suite. There, an advanced electron microscope will produce high-magnification images that detail important properties of the fly’s gut bacteria.
Ren Dodge carefully dissects a fly to prepare its gut for electron microscopy.
“We do a lot of experiments where we rip the fly apart,” says Ludington. “But that destroys its gut morphology. We knew that we also needed non-disruptive techniques that let us precisely quantify the geometry of the gut.”
So, Ludington has started to collaborate with the University of California’s Lawrence Berkeley National Lab, which uses a powerful X-ray machine called a synchrotron to produce detailed micro-CT scans of un-damaged flies.
Ludington Lab manager Daniel Martinez studies a micro-CT scan of a fly gut. The images to his right were generated at the Lawrence Berkeley National Lab’s Synchrotron facility.
“It's all about controlling variation. Everything we're doing is about controlling variation so that we know a precise perturbation causes a precise physiological reaction in the fly.”
Just two-and-a-half years after starting his lab at the Department of Embryology (the blink of an eye in biological science), Ludington has opened doors to many new possibilities for microbiome research and beyond—including the potential to help develop improved and novel treatments for human disease. He has already gained the attention and financial support of major scientific organizations, including the National Institution for Health (NIH), The National Science Foundation (NSF), and most recently, the Research Corporation For Science Advancement (RCSA). Stay tuned for updates!
Want to learn more? Visit The Ludington Lab