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Enhanced culturing techniques for the mycobiont isolated from the lichen Xanthoria parietina

ABSTRACT: Lichens and their isolated symbionts are potentially valuable resources for biotechnological approaches. Especially mycobiont cultures that produce secondary lichen products are receiving increasing attention, but lichen mycobionts are notoriously slow-growing organisms. Sufficient biomass production often represents a limiting factor for scientific and biotechnological investigations, requiring improvement of existing culturing techniques as well as methods for non-invasive assessment of growth. Here, the effects of pH and the supplement of growth media with either D-glucose or three different sugar alcohols that commonly occur in lichens, D-arabitol, D-mannitol and ribitol, on the growth of the axenically cultured mycobiont isolated from the lichen Xanthoria parietina were tested. Either D-glucose or different sugar alcohols were offered to the fungus at different concentrations, and cumulative growth and growth rates were assessed using two-dimensional image analysis over a period of 8 weeks. The mycobiont grew at a pH range from 4.0 to 7.0, whereas no growth was observed at higher pH values. Varying the carbon source in Lilly-Barnett medium (LBM) by replacing 1% D-glucose used in the originally described LBM by either 1%, 2% or 3% of D-mannitol, or 3% of D-glucose increased fungal biomass production by up to 26%, with an exponential growth phase between 2 and 6 weeks after inoculation. In summary, we present protocols for enhanced culture conditions and non-invasive assessment of growth of axenically cultured lichen mycobionts using image analysis, which may be useful for scientific and biotechnological approaches requiring cultured lichen mycobionts.

Supplementary Information

The online version contains supplementary material available at 10.1007/s11557-021-01707-7.

SUBMITTER: Pichler G

PROVIDER: S-EPMC8550697 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The postmortem microbiome has recently moved to the forefront of forensic research, and many studies have focused on the idea that predictable fluctuations in decomposer communities could be used as a "microbial clock" to determine time of death. Commonly, the oral microbiome has been evaluated using 16S rRNA gene sequencing to assess the changes in community composition throughout decomposition. We sampled the hard palates of three human donors over time to identify the prominent members of the microbiome. This study combined 16S rRNA sequencing with whole metagenomic (MetaG) and metatranscriptomic (MetaT) sequencing and culturing methodologies in an attempt to broaden current knowledge about how these postmortem microbiota change and might function throughout decomposition. In all four methods, Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes were the dominant phyla, but their distributions were insufficient in separating samples based on decomposition stage or time or by donor. Better resolution was observed at the level of genus, with fresher samples from decomposition clustering away from others via principal components analysis (PCA) of the sequencing data. Key genera in driving these trends included Rothia; Lysinibacillus, Lactobacillus, Staphylococcus, and other Firmicutes; and yeasts including Candida and Yarrowia. The majority of cultures (89%) matched to sequences obtained from at least one of the sequencing methods, while 11 cultures were found in the same samples using all three methods. These included Acinetobacter gerneri, Comamonas terrigena, Morganella morganii, Proteus vulgaris, Pseudomonas koreensis, Pseudomonas moraviensis, Raoutella terrigena, Stenotrophomonas maltophilia, Bacillus cereus, Kurthia zopfii, and Lactobacillus paracasei. MetaG and MetaT data also revealed many novel insects as likely visitors to the donors in this study, opening the door to investigating them as potential vectors of microorganisms during decomposition. The presence of cultures at specific time points in decomposition, including samples for which we have MetaT data, will yield future studies tying specific taxa to metabolic pathways involved in decomposition. Overall, we have shown that our 16S rRNA sequencing results from the human hard palate are consistent with other studies and have expanded on the range of taxa shown to be associated with human decomposition, including eukaryotes, based on additional sequencing technologies.

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Enhanced culturing techniques for the mycobiont isolated from the lichen Xanthoria parietina

Supplementary Information

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