Evolutionary Battleground: Plants vs. Microbes

Gazing out on a freshwater pond, you may see tiny green plants with oval shaped leaves floating in clusters. In overgrown ponds, these plants coat the water's surface. These plants - called duckweed or water lentils - can grow so fast that they can double their numbers in just one to two days. But what you can't see in that pond is the evolutionary battle between the plants and microbes trying to invade them.

Plants depend heavily on microbes around them. The community of bacteria, viruses, fungi and other microbes that accompany a plant is called a microbiome. Microbiomes are often specific to a type of plant in a particular location. There are often beneficial microbes that are part of a plant's microbiome. But at the same time, there are pathogenic microbes that the plants need to defend themselves against. Understanding how plants defend themselves against pathogens - or fail to do so - could help scientists know how to better manipulate microbiomes to benefit bioenergy crops.

Scientists supported by the Department of Energy's Office of Science are investigating how bacteria interact with plants' hormones and affect their growth.

An evolutionary showdown

One of plants' lines of defense against microbes is their stomata. Stomata are little pores on plants' leaves, stems, flowers, and roots that open and close to take in carbon dioxide and let out oxygen and water. They act like gates to a city. Just like gates, stomata create a physical barrier to invading bacteria. The plant has hormones that regulate whether the guard cells are keeping the stomata open or closed. However, certain bacteria can hack this system.

Another key player in this evolutionary back-and-forth are plant hormones called auxins. Auxins are an important class of hormones that affect how plants grow and develop. The most common auxin in nature is called Indole-3 acetic acid - IAA for short. In plants, IAA affects cells' length, plants' reaction to the direction of gravity, and the structure of roots. To protect against pathogens, plants can reduce IAA's impact. Because plants need to both grow and defend themselves, it's important for them to increase or decrease IAA when needed.

But as it so often happens in evolution, some bacteria have found a crack in this defense. Bacteria found in association with plants also produce IAA and do so in a similar way as plants. As part of that process, some of those bacteria evolved an override to the plants' IAA management. They produce enough IAA to affect the plants' chemical pathways and growth. Plants that have auxin pathways affected by these bacteria grow shorter primary roots than those that don't. They also grow more roots that run parallel to the surface of the ground and have more root hairs (the tiny root cells that bud from the surface layer of the root).

Peering into the process

These evolutionary fights are more than just an ecological wonder. They're also important for bioenergy crop productivity, agricultural productivity, and conservation. Charting how, when, and where bacteria produce plant hormones can help scientists understand how plants adapt to changing environments. Increasing the production of a specific hormone from a plant's microbiome or creating a synthetic microbiome could help farmers improve crop growth. Knowing how pathogens affect root growth and overall biomass could allow scientists to avert those problems when growing those plants to make biofuels.

To investigate this relationship, researchers from Rutgers University, the University of Tennessee, and the University of North Carolina at Chapel Hill studied duckweed. The genus of plants known as duckweed is extremely common and well-studied. Scientists have a lot of information about its genetics, including variations in DNA sequences between different populations of duckweed.

In a previous study, the group sampled the microbiome of duckweeds in the wild and found that it was similar to many common plants. Of 47 bacterial strains they analyzed, almost 80 percent produced compounds similar to IAA. With the support of scientists from the Environmental Molecular Sciences Laboratory (a DOE Office of Science user facility), the team inoculated duckweed seedlings with 21 of those strains. Some of the treated seedlings were regular "wild-type" in their root growth. Some plants had other bacteria strains that were producing the active auxin IAA. In these, the plants' roots grew much shorter and with much more branching. Similarly, plants missing a gene that is required to sense the presence of auxins were less sensitive to IAA than wild-type plants. These plants didn't respond to the IAA-producing bacteria.

Revealing the invasion

Surprisingly, of the 21 bacterial strains, only four affected plant growth in the way scientists expected. Not coincidentally, they were the ones that produced the most IAA in test tubes. It appears that although the rest could produce some compounds related to IAA, the amounts weren't enough to trigger a response from the plants. Those four seemed to be exceptionally good at getting past the plants' defenses.

But were those bacteria getting past the defenses because of the IAA or some other approach? The scientists answered that question by studying the mutant plants that lacked the gene that regulates auxin. They found that the bacteria could not simply move into the plant cells and colonize the mutant plants. Clearly, these bacteria had found a way to hack the auxin system to work in their favor.

The next question was where the bacteria got in. While plants' stomata are a defense mechanism, they're also a weakness. Using special dyes and microscopes, the researchers found loads of the invading bacteria in the stomata of the wild-type duckweed. In contrast, most of the bacteria in the mutant plants were on the surface, where they couldn't home in on the stomata. Like an invader tricking the guards at the gates of a walled city, the successful bacteria used the auxin system to trick the guard cells.

Tracking just a little of this evolutionary tug-of-war between plants and bacteria suggests that plants are likely to have multiple defense mechanisms against bacteria that produce IAA.

While the leaves and stems of plants can be readily seen above ground, a whole complex system of cooperative and competitive behavior is hidden from view. By illuminating these relationships, scientists are setting the foundation to grow plants for biofuels in more sustainable ways.