Thallus Growth Stage and Geographic Origin Shape Microalgal Diversity in Ramalina farinacea Lichen Holobionts

Abstract

Lichen symbioses are microecosystems hosting many other living organisms besides the two major lichen symbionts (i.e., lichenized fungi [the mycobiont] and green microalgae or cyanobacteria [the photobiont]). Recent investigations evidenced that other fungi, non‐photosynthetic bacteria, and microalgae co‐inhabit within the lichen thalli, but their diversity and their roles are still underinvestigated. Here we present an ad hoc stratified sampling design and in‐depth Illumina paired‐end metabarcoding approach to explore microalgal diversity in lichen thalli of the model species Ramalina farinacea from different ecologies. Lichen thalli were surveyed according to three different sizes, and different thallus parts were considered for molecular, bioinformatics, and community diversity analyses. The results revealed that microalgal diversity strongly depends on the growth stage of the thalli, the geographic area, and the habitat type. The results also show that microalgal diversity does not vary along the thallus branches (lacinias)—that is, it does not correlate with the apical growth and founder effects—and that there is no balanced co‐presence of two main photobionts as previously established in R. farinacea. The sampling design performed here minimizes bias in the assessment of photobiont diversity in lichens and is proposed to be reliable and applicable to further study microalgal diversity in lichen symbioses.

The Role of Photobionts as Drivers of Diversification in an Island Radiation of Lichen-Forming Fungi

“…Thallus tips were the only zones free of epiphytes in all samples. The choice of thallus tips is also supported by the finding of Molins et al (2021) , who reported that there were no differences in photobiont composition among different parts of the thallus in specimens of a species of the same genus ( R. farinacea ) from the same biogeographic region as the present study. Samples were washed with acetone to remove secondary metabolites and stored at −80°C.…”

“…TR9 as main photobiont is not exclusive of the R. decipiens group. It was found in Macaronesian specimens of R. farinacea using Sanger sequencing ( del Campo et al, 2013 ) and NGS ( Molins et al, 2021 ). BLAST searches of the two predominant ASVs revealed their presence also in other genera.…”

Watanabea green microalgae (Trebouxiophyceae) inhabiting lichen holobiomes: Watanabea lichenicola sp. nova

“…As in previous studies (Chiva 2020;Molins et al 2020;Moya et al 2020), the ultrastructure of symbiotic microalgae observed in lichen fragment of both thalli (MT27 and TE1) fits with typical features of the genus Trebouxia (Appendix S4 in the Supporting Information). In studies carried out on the diversity of microalgae in lichen thalli by massive sequencing (454, Illumina), several genera of green microalgae were always detected (Moya et al 2017;Molins et al 2021), although in a low proportion and, in general, their sequences are not usually available in GenBank. Moreover, no microalgae have been identified with the genetic and morphological characteristics of the genus Watanabea.…”

“…Lichen thalli are complex symbiotic micro-ecosystems where an exhabitant fungus (mycobiont) interacts with extracellular lineages of microalgae (phycobionts/photobionts). Additionally, non-photosynthetic bacteria, other fungi and diverse green microalgae also participate in the symbiosis, self-maintaining the viability of the thalli (holobionts) (Aschenbrenner et al 2016;Smith et al 2020;Molins et al 2021). The symbiogenetic phenotypes resulting from mutualistic interactions within microbial assemblages provide a survival strategy to thrive under extreme conditions, for example in polar areas and xeric deserts (Barreno 2013;Raggio et al 2021).…”

“…Likewise, these multipartite interactions are the scenario of poorly known interaction networks, as well as being a source of biodiversity and new species (Krug et al 2020). Among these potential associated organisms, diverse green coccoid algae could be considered as part of the lichen phycobiomes (Moya et al 2017;Molins et al 2021 (Muggia et al 2020). This means increased knowledge concerning biodiversity, and an undoubted potential for emerging new organisms to be used in systematic, phylogenetic, ecological or biotechnological studies.…”