So it turns out to be a threesome

Lichens are amazing organisms, drawing out our curiosity both because of their aesthetic appeal and their novel ecological and evolutionary characteristics. For over a hundred years, biologists have supposedly known about their basic composition: they are one of nature’s best examples of a symbiosis, a tight coupling of two or more evolutionarily distinct organisms that can function like a single unit. In the case of lichens, the two coupled organisms are a fungus and a photosynthetic alga.

Encrusting lichens at Newberry National Monument, Deschutes Co., OR, June 2011.

That basic understanding of lichens as an evolutionary marriage between alga and fungus remains fact. However, a paper published this summer significantly expanded the overall picture. Lead author Toby Spribille and his colleagues provide evidence in the journal Science that many lichens are symbioses of not just two groups of organisms, but three! How could such a basic aspect of lichen biology go undetected for so long?

Letharia vulpina on tree bark at Diamond Peak
Wilderness, OR, May 2013.

Their surprising discovery came about by careful observation and testing of what might otherwise pass as a rather mundane question in lichen biology. The authors were investigating differences in two closely related species of lichen, Bryoria tortuosa and B. fremontii. The former species produces a compound called vulpinic acid that causes the lichen to have a yellowish color. B. fremontii, however, lacks (or has reduced production of) the acid and is brown in color. What had baffled researchers, however, was the failure to find genetic differences between the two supposed species. Both the ascomycete fungus and the photosynthetic partner (a green alga called Trebouxia simplex) in the two species had identical sequences when several genes were studied. Species are expected to have fixed genetic differences even if their phenotype (their appearance) is very similar.

Spribille and colleagues decided to study the genetic structure again of the two species by sequencing the mRNA transcriptome. They confirmed the genetic similarity that had been observed before and also found little difference in gene expression between B. tortuosa and B. fremontii. However, when the researchers broadened their search to consider whether the transcripts they sequenced might match any other types of fungi, they found that the acid-producing B. tortuosa produced sequences associated with basidiomycetes, a very different group of fungi than are typically seen in lichens. The evidence pointed to a third symbiont in B. tortousa!

Apothecia (spore-producing structures) on encrusting lichens. Left: Alpine bloodspot, Ophioparma ventosa, Deschutes National Forest, OR, May 2012. Right: Ochrolechia sp., Bear Valley, Tahoe National Forest, CA, Dec 2009.

Usnea longissima, hanging from a vine maple in the
Menagerie Wilderness, Linn Co., OR, Oct 2012.

With this new finding, the researchers asked whether other lichen species might potentially harbor basidiomycetes as well. Surveying a variety of other lichen lineages, they found basidiomycete sequences in 52 different genera distributed across six continents! The basidiomycete lineages in the lichens seemed to be diverse group of fungi, but they were associated with specific species. The new group of basidiomycete symbionts is called the Cyphobasidiales.

Up until now, it had been known that some lichens deviated from the classic model of two partners in the lichen symbiosis. For instance, some species have two algal hosts in addition to the fungal partner – both a green alga and a cyanobacterium (Henskens et al. 2012). However, Spribille et al.’s research suggests that the typical lichen association is made up of two distinct fungal lineages and one or two algae. Thus, lichens may typically be threesomes, and sometimes, even foursomes. As if this wasn’t complicated enough, DNA sequencing work by Bates et al. (2011) showed that lichens can also have bacterial communities associated with them. They found unique groups of alpha proteobacteria associated with the lichen body in 4 species they studied. Apparently the more that lichens are studied, the more we could describe these remarkable organisms as comprising their own little ecological worlds!

The lichen Cladonia in a residential yard, Humboldt
Co., CA, Dec 2009.

With the relative ubiquity of the third symbiont evident in sequence data, why haven’t previous researchers seen these basidiomycete symbionts when looking through the microscope at lichen specimens? It turns out that the cells are difficult to detect by microscopy. They are small, and on the periphery of the lichen body, being embedded in a matrix of polysaccharides. Using florescent molecular tags on rRNA sequences specific to the basidiomycete partner, however, the new cells and their location in the lichen thallus (body) became readily apparent.

Interestingly, the new results explain one of the mysteries that lichen biologists have confronted for a long time, namely that it has been difficult to recreate the lichen symbiosis in the lab by combining only a single fungal host and alga. The typical lichen shape was hard to reproduce with only two partners.

It is fascinating to ponder the evolutionary history that could lead to such a complex and intimate association of distinct organisms. Green algae, cyanobacteria, and fungi are not closely related. During the early evolution of lichens, how did algae first become associated with the body of fungi? Did early lichens start with two fungal partners plus an alga, or were today’s lichen ancestors more simple in composition?

Lichens covering a rock in coastal scrub near
Muir Beach, Marin Co., CA, Oct 2016.

Whatever the exact evolutionary sequence of events, the lichen symbiosis appears to have proved to be very successful in terms of survival and reproductive success. In fact, fossils suggest that associations of algae and fungi are at least a half billion years old, stretching back to about the time that invertebrate animals diversified in the Cambrian Explosion, and before the arrival of vascular plants on land (Yuan et al. 2005). Today, there are many thousands of species of lichens, inhabiting ecosystems as diverse as deserts, coniferous rainforests, and coastlines.

References

Bates ST, Cropsey GW, Caporaso JG, Knight R, Fierer N. 2011. Bacterial communities associated with the lichen symbiosis. Applied and Environmental Microbiology 77:1309-1314.

Brodo IM, Sharnoff SD, Sharnoff S. 2001. Lichens of North America. Yale University Press.

Henskens FL, Green TG, Wilkins A. 2012. Cyanolichens can have both cyanobacteria and green algae in a common layer as major contributors to photosynthesis. Annals of Botany 110:555-563.

Spribille T, Touvinen V, Resl P, et al. 2016. Basidiomycete yeasts in the cortex of ascomycete lichens. Science 353:488-492.

Yuan X, Xiao S, Taylor TN. 2005. Lichen-like symbiosis 600 million years ago. Science 308:1017-1020.

Lichen (perhaps Amandinea) on wood fence, Pt. Reyes National Seashore, CA, Nov 2008.

Fruticose lichen in redwood forest, Van Duzen County Park, Humboldt Co., CA, 2006.