Hidden allies

Endophytes: A plant’s friends or foes

Endophytes are microorganisms that live inside plants. Some of these organisms, mostly bacteria or fungi, make the plants sick, while others have no harmful effect on the plants or are even beneficial. Previous studies of endophytic fungi living inside certain grasses have shown that these fungi provide the grasses with a defense against predators. However, little has been known about whether this is also true for trees. The current study investigated the influence of an endophytic fungus of the genus Cladosporium on the herbivore defense of the black poplar Populus nigra, as well as the effects on the insect communities that live on poplars.

"We investigated the influence of the endophyte on the chemical defense of black poplars and the consequences for the food preference and fitness of herbivorous insects, as well as the composition of the insect community on young trees in the field. We already knew from previous studies that plant pathogenic rust fungi are beneficial to herbivorous insects. Now we were interested in an endophytic fungus whose role in poplar-insect interactions is still largely unexplored," says first author Christin Walther, explaining the starting point of the study.

The researchers found that colonization with the fungal endophyte boosted the poplars' natural defenses and those induced by insect damage. In laboratory experiments, the larvae of the gypsy moth Lymantria dispar, a generalist on various trees, preferred uninfected leaves and performed better there. "We were able to show that Cladosporium helps the plant to defend itself against predators. To do this, the fungus alters the plant's chemical defense profile and also supports it with a self-produced defense substance," says Christin Walther.

Using the state-of-the-art chemical analysis method known as metabolomics, which analyses the metabolic products in the leaves, the scientists were able to show that infection with the fungus changes the poplar metabolome, i.e. the substances produced in the poplar. Specific defense substances such as salicinoids (salicin, nigracin) and phenolic acids (caffeic acid) were produced at higher levels in the presence of the fungus. Together with tannins, salicinoids are the most important natural defense substances produced by poplars to protect themselves. Salicinoids are particularly effective against generalist insects that are not only specialized on poplar, but are also pests of other plants. As well as deterring pests, they also have a negative effect on the fitness of insects that feed on chemically defended leaves.

Cladosporium defends poplars with its own defense substance

The analysis revealed that the fungus itself produces an effective defense compound, the alkaloid stachydrine. Tests with stachydrine showed that the endophyte's active substance fended off both generalists, such as the brush moth Orgyia antiqua, and poplar specialists, such as the poplar leaf beetle Chrysomela tremulae.

 "It was particularly exciting for us to see that the fungus can produce an alkaloid that we can detect in the plant matrix and thus use as a marker for the presence of the fungus. We even found this substance in the leaves of old trees in the field," says study leader Sybille Unsicker, who headed the "Plant-Environment Interactions" group at the Max Planck Institute for Chemical Ecology and has been a professor at Kiel University since 2023.

Combination of chemical analyses, behavioral experiments and field studies

The scientists took a holistic approach to their research. In addition to chemical analyses of laboratory plants with and without the endophytic fungus and associated feeding experiments, they investigated in the field how Cladosporium affects the chemical ecology of black poplars.

Another interesting result of this study is an observation that the researchers made in the field. The endophytic fungus also seems to alter the interactions between aphids and ants that colonize young black poplars. The ant-aphid-relationship is a mutualism, a symbiosis that benefits both the ants, who drink the aphids' honeydew, and the aphids, who are protected from predators by the ants. Plants that had previously been infected with the endophyte were more often colonized by aphids, but less often by ants. This is probably due to the concentration of stachydrine in their honeydew. "The fungus even seems to be able to manipulate the mutualistic relationship between ants and aphids. Further research is needed to clarify exactly what is happening," says Christin Walther.

Trees are metaorganisms

Studying a tree's microbiome, the entirety of the microorganisms associated with the tree, is important for understanding the complexity of the co-evolution of trees and other organisms with which they interact. The current study is just the beginning of a broad field of research that raises many other exciting questions. For example, further studies will look at how the fungus spreads through the plant and whether it protects the plant not only from insect damage, but possibly also from pathogens. Another interesting question is whether the relationship between black poplar and Cladosporium is symbiotic, and whether the fungus benefits from living with the tree, or whether there are conditions under which it turns from a partner to a threat to the tree.

"There is a growing body of evidence in favor of considering trees as holobionts, as communities of different organisms. A tree is never a single individual, but a complex metaorganism colonized by microorganisms and arthropods in all its organs. The exciting question for us is how microorganisms contribute to the co-evolution of trees and herbivorous insects. Our results are certainly only a small contribution to understanding these complex interactions," concludes Sybille Unsicker.