Scientists have discovered a previously unidentified bacterium in the termite gut that may play a key role in turning hard, fibrous material into a nutritious meal.
When termites munch on wood, the small bits are delivered to feed a community of unique microbes living in their guts, and in a complex process involving multiple steps, these microbes process the material for the termite host.
One key step uses hydrogen to convert carbon dioxide into organic carbon—a process called acetogenesis—but little is known about which gut bacteria play specific roles in the process.
“In the termite gut, you have several hundred different species of microbes that live within a millimeter of one another,” says Jared Leadbetter, professor of environmental microbiology at California Institute of Technology (Caltech).
“We know certain microbes are present in the gut, and we know microbes are responsible for certain functions, but until now, we didn’t have a good way of knowing which microbes are doing what.”
Acetogenesis is the production of acetate (a source of nutrition for termites) from the carbon dioxide and hydrogen generated by gut protozoa as they break down decaying wood.
Total gene pool
In their study, published in Proceedings of the National Academy of Sciences, researchers searched the entire pool of termite gut microbes to identify specific genes from organisms responsible for acetogenesis.
They began by sifting through the microbes’ RNA—genetic information that can provide a snapshot of the genes active at a certain point in time. Using RNA from the total pool of termite gut microbes, they searched for actively transcribed formate dehydrogenase (FDH) genes, known to encode a protein necessary for acetogenesis.
Next, using a method called multiplex microfluidic digital polymerase chain reaction (digital PCR), researchers sequestered the previously unstudied individual microbes into tiny compartments to identify the actual microbial species carrying each of the FDH genes.
Some of the FDH genes were found in types of bacteria known as spirochetes—a previously predicted source of acetogenesis. Yet it appeared that these spirochetes alone could not account for all of the acetate produced in the termite gut.
Initially, the researchers were unable to identify the microorganism expressing the single most active FDH gene in the gut. However, the first authors on the study, Adam Rosenthal, a postdoctoral scholar in biology and Xinning Zhang, noticed that this gene was more abundant in the portion of the gut extract containing wood chunks and larger microbes, like protozoans.
Chunky gut stuff
After analyzing the chunkier gut extract, they discovered that the single most active FDH gene was encoded by a previously unstudied species from a group of microbes known as the deltaproteobacteria. This was the first evidence that a substantial amount of acetate in the gut may be produced by a non-spirochete.
Because the genes from this deltaproteobacterium were found in the chunky particulate matter of the termite gut, the researchers thought that perhaps the newly identified microbe attaches to the surface of one of the chunks. To test this hypothesis, they used a color-coded visualization method called hybridization chain reaction-fluorescent in situ hybridization, or HCR-FISH.
The technique allowed the researchers to simultaneously “paint” cells expressing both the active FDH gene and a gene identifying the deltoproteobacterium with different fluorescent colors simultaneously.
“The microfluidics experiment suggested that the two colors should be expressed in the same location and in the same tiny cell,” Leadbetter says. And, indeed, they were.
“Through this approach, we were able to actually see where the new deltaproteobacterium resided. As it turns out, the cells live on the surface of a very particular hydrogen-producing protozoan.”
This association between the two organisms makes sense based on what is known about the complex food web of the termite gut, Leadbetter says.
“Here you have a large eukaryotic single cell—a protozoan—which is making hydrogen as it degrades wood, and you have these much smaller hydrogen-consuming deltaproteobacteria attached to its surface,” he says.
“So, this new acetogenic bacterium is snuggled up to its source of hydrogen just as close as it can get.”
This intimate relationship might never have been discovered relying on phylogenetic inference—the standard method for matching a function to a specific organism.
“Using phylogenetic inference, we say, ‘We know a lot about this hypothetical organism’s relatives, so without ever seeing the organism, we’re going to make guesses about who it is related to,” he says.
“But with the techniques in this study, we found that our initial prediction was wrong. Importantly, we have been able to determine the specific organism responsible and a location of the mystery organism, both of which appear to be extremely important in the consumption of hydrogen and turning it into a product the insect can use.”
The results not only identify a new source for acetogenesis in the termite gut, the researchers say, they also reveal the limitations of making predictions based exclusively on phylogenetic relationships.
The US Department of Energy, the National Science Foundation, the National Institutes of Health, the Programmable Molecular Technology Center within the Beckman Institute at Caltech, a Donna and Benjamin M. Rosen Center Bioengineering scholarship, and the Center for Environmental Microbial Interactions at Caltech supported the work.