TY - JOUR

T1 - Unraveling intragenomic polymorphisms in the high-quality genome of Hypoxylaceae: a comprehensive study of the rDNA cistron

AU - Cedeño-Sanchez, Marjorie

AU - Cheng, Tian

AU - Lambert, Christopher

AU - Kolarík, Miroslav

AU - Kuhnert, Eric

AU - Cox, Russell J.

AU - Kalinowski, Jörn

AU - Verwaaijen, Bart

AU - Stadler, Marc

N1 - Funding Information: Open Access funding enabled and organized by Projekt DEAL. This work was funded by the DFG (Deutsche Forschungsgemeinschaft) priority program “Taxon-Omics: New Approaches for Discovering and Naming Biodiversity” (SPP 1991), specifically CO 1328/4-2 and CO 1328/4-1 and the European Union’s H2020 Research and Innovation Staff Exchange program (MSCA-RISE grant no. 101008129, Acronym: MYCOBIOMICS). The bioinformatics support of the BMBF-funded project “Bielefeld-Gießen Center for Microbial Bioinformatics; BiGi (grant no. 031A533)” within the German Network for Bioinformatics Infrastructure (de.NBI) is gratefully acknowledged. MCS gratefully acknowledges a PhD stipend from The National Secretariat of Science, Technology and Innovation of the Republic of Panama (SENACYT) and the Institute for the Development of Human Resources (IFARHU). CL is grateful for financial support in form of a stipend from the Life-Science Foundation (LSS Munich).

PY - 2024

Y1 - 2024

N2 - Sequencing ribosomal DNA loci, such as ITS and LSU, are among the first steps to characterize and identify newly isolated fungal strains. Substantial doubt has recently been raised concerning the unequivocal suitability of using this sequence information for inferring inter- and intraspecific relationships, as especially the ITS locus was repeatedly shown to harbor high intragenomic plasticity. The evolutionary mechanisms leading to this phenomenon are poorly understood, but this is about to change, owing to the availability of third generation sequencing techniques. The current study attempts a comprehensive analysis of rDNA cistrons in the genomes of over 40 selected species of Hypoxylaceae (Xylariales), using a combination of 2nd generation (Illumina) and 3rd generation sequencing technologies (Oxford Nanopore), along with extensive usage of bioinformatic tools to obtain high-quality genomes. Our findings revealed high intragenomic polymorphisms in distinct Hypoxylaceae species, indicating the presence of deep rDNA paralogs. Phylogenetic inference of the ITS rDNA showed low support for the primary clades within Hypoxylaceae, while inference using the protein encoding gene RPB2 showed better support. We also found that proteinogenic genes are not prone to intraspecific and intragenomic polymorphisms as much as the rDNA loci. Our study highlights the limitations of solely relying on the rDNA cistron as a barcode for fungi. We propose TUB2 as a new primary barcoding marker for Hypoxylaceae and other Xylariales. Moreover, we demonstrate the feasibility to retrieve some DNA loci that had hitherto not been used for classical phylogenetic studies (such as TEF1) in Xylariales directly from genomes without the necessity to work with primers.

AB - Sequencing ribosomal DNA loci, such as ITS and LSU, are among the first steps to characterize and identify newly isolated fungal strains. Substantial doubt has recently been raised concerning the unequivocal suitability of using this sequence information for inferring inter- and intraspecific relationships, as especially the ITS locus was repeatedly shown to harbor high intragenomic plasticity. The evolutionary mechanisms leading to this phenomenon are poorly understood, but this is about to change, owing to the availability of third generation sequencing techniques. The current study attempts a comprehensive analysis of rDNA cistrons in the genomes of over 40 selected species of Hypoxylaceae (Xylariales), using a combination of 2nd generation (Illumina) and 3rd generation sequencing technologies (Oxford Nanopore), along with extensive usage of bioinformatic tools to obtain high-quality genomes. Our findings revealed high intragenomic polymorphisms in distinct Hypoxylaceae species, indicating the presence of deep rDNA paralogs. Phylogenetic inference of the ITS rDNA showed low support for the primary clades within Hypoxylaceae, while inference using the protein encoding gene RPB2 showed better support. We also found that proteinogenic genes are not prone to intraspecific and intragenomic polymorphisms as much as the rDNA loci. Our study highlights the limitations of solely relying on the rDNA cistron as a barcode for fungi. We propose TUB2 as a new primary barcoding marker for Hypoxylaceae and other Xylariales. Moreover, we demonstrate the feasibility to retrieve some DNA loci that had hitherto not been used for classical phylogenetic studies (such as TEF1) in Xylariales directly from genomes without the necessity to work with primers.

KW - Barcoding

KW - Genomics

KW - Phylogeny

KW - Sanger sequencing

KW - Xylariales

UR - http://www.scopus.com/inward/record.url?scp=85180712530&partnerID=8YFLogxK

U2 - 10.1007/s11557-023-01940-2

DO - 10.1007/s11557-023-01940-2

M3 - Article

AN - SCOPUS:85180712530

VL - 23

JO - Mycological progress

JF - Mycological progress

SN - 1617-416X

IS - 1

M1 - 5

ER -