Dataset Information

Variability in ITS1 and ITS2 sequences of historic herbaria and extant (fresh) Phalaris species (Poaceae)

ABSTRACT:

Background

Phalaris species (Poaceae) occupy diverse environments throughout all continents except Antarctica. Phalaris arundinacea is an important forage, ornamental, wetland restoration and biofuel crop grown globally as well as being a wetland invasive. The nuclear ribosomal internal transcribed spacer (ITS) region has been used for Phalaris barcoding as a DNA region with high nucleotide diversity for Phalaris species identification. Recent findings that P. arundinacea populations in Minnesota USA are most likely native and not European prompted this analysis to determine whether Eurasian vs. native North American P. arundinacea differed in ITS regions. Our objectives were to amplify and compare ITS regions (ITS1 and ITS2) of historic herbaria (1882–2001) and extant (fresh) Phalaris specimens; analyze ITS regions for species-specific polymorphisms (diagnostic SNPs) and compare ITS regions of historic Phalaris specimens with known, extant Phalaris species.

Results

We obtained complete ITS1 and ITS2 sequences from 31 Phalaris historic (herbaria samples, 1908 to 2001) and five extant (fresh) specimens. Herbaria Phalaris specimens did not produce new SNPs (single nucleotide polymorphisms) not present in extant specimens. Diagnostic SNPs were identified in 8/12 (66.6%) Phalaris species. This study demonstrates the use of herbaria tissue for barcoding as a means for improved species identification of Phalaris herbaria specimens. No significant correlation between specimen age and genomic DNA concentration was found. Phalaris arundinacea showed high SNP variation within its clade, with the North American being distinctly different than other USA and most Eurasian types, potentially allowing for future identification of specific SNPs to geographic origin.

Conclusions

While not as efficient as extant specimens to obtain DNA, Phalaris herbaria specimens can produce high quality ITS sequences to evaluate historic genetic resources and facilitate identification of new species-specific barcodes. No correlation between DNA concentration and age of historic samples (119 year range) occurred. Considerable polymorphism was exhibited in the P. arundinacea clade with several N. American accessions being distinct from Eurasian types. Further development of within species- and genus-specific barcodes could contribute to designing PCR primers for efficient and accurate identification of N. American P. arundinacea. Our finding of misidentified Phalaris species indicates the need to exercise stringent quality control measures on newly generated sequence data and to approach public sequence databases in a critical way.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12870-021-03284-z.

SUBMITTER: Graper A

PROVIDER: S-EPMC8571858 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:This paper presents the performance of two eukaryotic genomic ribosomal regions, ITS1 and ITS2, in describing fungal diversity in aerosol samples using amplicon-based High-Throughput Sequencing (HTS). Composting sites, biomethanization facilities, and dairy farms, all affected by the presence of fungi, were visited to collect air samples. The amplicon-based HTS approach is a target enrichment method that relies on the amplification of a specific target using particular primers before sequencing. Thus, the results are highly dependent on the quality of amplification. For this reason, the authors of this paper used a shotgun metagenomic approach to compare its outcome with the amplicon-based method. Indeed, shotgun metagenomic does not rely on any amplification prior to sequencing, because all genes are sequenced without a specific target. In addition, culture methods were added to the analyses in biomethanization and dairy farms samples to validate their contribution to fungal diversity of aerosols. The results obtained are unequivocal towards ITS1 outperformance to ITS2 in terms of richness, and taxonomic coverage. The differential abundance analysis did demonstrate that some taxa were exclusively detected only by ITS2, and vice-versa for ITS1. However, the shotgun metagenomic approach showed a taxonomic profile more resembling to ITS1 than ITS2. Based on these results, neither of the barcodes evaluated is perfect in terms of distinguishing all species. Using both barcodes offers a broader view of the fungal aerosol population. However, with the actual knowledge, the authors strongly recommend using ITS1 as a universal fungal barcode for quick general analyses of diversity and when limited financial resources are available, primarily due its ability to capture taxonomic profiles similar to those obtained using the shotgun metagenomic. The culture comparison with amplicon-based sequencing showed the complementarity of both approaches in describing the most abundant taxa.