Abstract
The implementation of HTS (high-throughput sequencing) approaches is rapidly changing our understanding of the lichen symbiosis, by uncovering high bacterial and fungal diversity, which is often host-specific. Recently, HTS methods revealed the presence of multiple photobionts inside a single thallus in several lichen species. This differs from Sanger technology, which typically yields a single, unambiguous algal sequence per individual. Here we compared HTS and Sanger methods for estimating the diversity of green algal symbionts within lichen thalli using 240 lichen individuals belonging to two species of lichen-forming fungi. According to HTS data, Sanger technology consistently yielded the most abundant photobiont sequence in the sample. However, if the second most abundant photobiont exceeded 30% of the total HTS reads in a sample, Sanger sequencing generally failed. Our results suggest that most lichen individuals in the two analyzed species, Lasallia hispanica and L. pustulata, indeed contain a single, predominant green algal photobiont. We conclude that Sanger sequencing is a valid approach to detect the dominant photobionts in lichen individuals and populations. We discuss which research areas in lichen ecology and evolution will continue to benefit from Sanger sequencing, and which areas will profit from HTS approaches to assessing symbiont diversity.
Highlights
The implementation of High-throughput sequencing (HTS) approaches is rapidly changing our understanding of the lichen symbiosis, by uncovering high bacterial and fungal diversity, which is often host-specific
Recent HTS studies on lichen photobiont diversity suggest that the presence of multiple, genetically differentiated algae within a single thallus is a common phenomenon in lichens[24,25,26]
We found no differences in diversity between ITS1 and ITS2 for the Trebouxia algae associated with the two lichen species, Lasallia pustulata and L. hispanica
Summary
The implementation of HTS (high-throughput sequencing) approaches is rapidly changing our understanding of the lichen symbiosis, by uncovering high bacterial and fungal diversity, which is often host-specific. Because HTS of DNA barcodes, e.g. on the Illumina MiSeq, Ion Torrent, or PacBio platforms, allows taxonomic assessment at a far greater depth and resolution than conventional Sanger sequencing[2,7], it facilitates discovery of rare taxa and detection of previously unrecognized eukaryotic and prokaryotic microbiomes[8,9,10,11,12] Studying this vastly unexplored microbial biosphere is one of the new frontiers of microbial ecology, currently changing our understanding of species interactions and their ecological and evolutionary dynamics[13]. Recent HTS studies on lichen photobiont diversity suggest that the presence of multiple, genetically differentiated algae within a single thallus is a common phenomenon in lichens[24,25,26] This raises the question to what extent Sanger-based studies underestimate within-thallus green algal diversity, and whether the single Sanger sequence obtained from an individual corresponds to the most abundant photobiont in that individual[27].
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