Deciphering functional diversification within the lichen microbiota by meta-omics

Abstract

Background: Recent evidence of specific bacterial communities extended the traditional concept of fungal-algal lichen symbioses by a further organismal kingdom. Although functional roles were already assigned to dominant members of the highly diversified microbiota, a substantial fraction of the ubiquitous colonizers remained unexplored. We employed a multi-omics approach to further characterize functional guilds in an unconventional model system.

Results: The general community structure of the lichen-associated microbiota was shown to be highly similar irrespective of the employed omics approach. Five highly abundant bacterial orders-Sphingomonadales, Rhodospirillales, Myxococcales, Chthoniobacterales, and Sphingobacteriales-harbor functions that are of substantial importance for the holobiome. Identified functions range from the provision of vitamins and cofactors to the degradation of phenolic compounds like phenylpropanoid, xylenols, and cresols.

Conclusions: Functions that facilitate the persistence of Lobaria pulmonaria under unfavorable conditions were present in previously overlooked fractions of the microbiota. So far, unrecognized groups like Chthoniobacterales (Verrucomicrobia) emerged as functional protectors in the lichen microbiome. By combining multi-omics and imaging techniques, we highlight previously overlooked participants in the complex microenvironment of the lichens.

Keywords: Amplicon sequencing; Lichen symbiosis; Lobaria pulmonaria; Metagenomics; Metaproteomics; Metatranscriptomics.

Fig. 1

Bacterial community structures of the Lobaria-associated microbiome are visualized in a mulit-level Krona chart from phylum to order level (starting at the inner circle) for each omic approach (16S rNA amplicons, metagenome, metatranscriptome). Taxonomic information is based on the 16S rRNA gene fragment sequence analysis performed with QIIME

Fig. 2

Relative sequence abundances of families within the highly abundant orders Sphingobacteriales, Rhodospirillales, Sphingomonadales, Myxococcales, and Chthoniobacterales according to the three omic approaches (16S rRNA amplicon, metagenome, and metatranscriptome). Unassigned or unclassified families were grouped into “others”

Fig. 3

Functional assignments to selected orders within the Lobaria-associated microbiome based on the metagenome, transcriptome, and proteome. Functional information of Chthoniobacterales, Myxococcales, Rhodospirillales, Sphingomonadales, and Sphingobacteriales based on the transcriptome and genome was obtained by blastx assignment, followed by processing with MEGAN including SEED classification (a). The COG database was utilized for functional classification of the proteome data (b)

Fig. 4

Colonization patterns of Spartobacteria (a, b) and Verrucomicrobia (c) on the lichen thallus of L. pulmonaria stained by fluorescence in situ hybridization (FISH). Green: algae Dictyochloropsis reticulata; gray or blue/purple: lichenized fungus L. pulmonaria; yellow: Spartobacteria or Verrucomicrobia; red, other eubacteria. a, b Volume rendering of confocal stacks. c Three-dimensional model reconstruction visualized as isosurface and spheres. Arrows or circles indicate single colonies or areas with small clusters of Spartobacteria or Verrucomicrobia within unspecific-labeled bacterial aggregates

Fig. 5

Functional assignments within the lichen symbiosis obtained with multi-omics technologies and corresponding analyses. Bacterial groups are shown on different taxonomic levels. Colored bubbles include results from the current study, while gray bubbles were adapted with data from Cernava et al. [15, 16], Erlacher et al. [22], and Grube et al. [28]. Bubble sizes correspond to the mean abundance of each group, such with a rectangular label field are present with <1% in the Lobaria microbiome