What Quality Suffices for Nanopore Metabarcoding? Reconsidering Methodology and Ectomycorrhizae in Decaying Fagus sylvatica Bark as Case Study
Abstract
:1. Introduction
1.1. Fungal Metabarcoding
1.2. Case Study: Deadwood Bark Ectomycorrhizae
2. Materials and Methods
2.1. Study Site and Log Selection
2.2. Root Tip Morphotypification and Sanger Barcoding
2.3. Nanopore Metabarcoding
2.3.1. Mock Community
2.3.2. Bark Sampling
2.3.3. Wet Lab
2.4. Bioinformatics
2.4.1. Reference Database
2.4.2. eNano Pipeline
2.4.3. SH Approach
2.4.4. OTU Approach
2.4.5. Minimum Quality Evaluation
2.4.6. Bark Metabarcoding
3. Results
3.1. Morphotyping
3.2. Metabarcoding
3.2.1. Minimum Quality Evaluation
3.2.2. Mock Community
3.2.3. Bark Substrate Community
3.2.4. Mycorrhizae in Decaying Beech Inner Bark
4. Discussion
4.1. Minimum Quality Evaluation
4.2. Mock Community
4.3. Inner Bark Community
4.4. Ectomycorrhizal Colonization of Beech Deadwood
4.5. Database
4.6. Nanopore Metabarcoding
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Bark Decay Stage | Description |
---|---|
Early | Layer (incl. inner bark) is difficult to penetrate with a metal spoon (force needed), and almost all fibers are intact and dark yellow in color. This corresponds to log decay stage (DS) 1 or early 2. |
Intermediate | Layer crumbles and can be scooped up with a metal spoon. They crumble with minimal force between fingers, some fibers intact, light brown in color. This corresponds to a late log DS 2 or early 3. |
Late | Layer is almost fully decayed, penetrable without force; it easily disintegrates between fingertips–resembles soil, no fibers intact, (dark) brown in color. Corresponds to a late log DS 3 or 4. |
DS Bark | Lowest Taxon Name | Fidelity | p | SH|OTU | |
---|---|---|---|---|---|
Early | Chaetosphaeriaceae (SH0980871.10FU) | 0.798 | 0.001 | ● | ● |
Tulasnella (OTU 1097) | 0.728 | 0.003 | ● | ||
Sordariales (SH0840221.10FU) | 0.779 | 0.008 | ● | ● | |
Pleurothecium recurvatum (SH0926211.10FU) * | 0.950 | 0.007 | ● | ● | |
Mid | Ganoderma adspersum (SH0762773.10FU) | 0.839 | 0.001 | ● | |
Hydnodontaceae (OTU 448) | 0.729 | 0.004 | ● | ||
Herpotrichiellaceae (SH0970954.10FU) | 0.751 | 0.005 | ● | ● | |
Rhinocladiella (OTU 75|SH0970950.10FU) | 0.697 | 0.005 | ● | ||
Helotiales (OTU 474) | 0.729 | 0.006 | ● | ||
Mortierella (SH0960682.10FU) | 0.745 | 0.008 | ● | ● | |
Serendipita (SH0743656.10FU) | 0.730 | 0.008 | ● | ● | |
Chaetosphaeria decastyla (SH0980872.10FU) | 0.655 | 0.010 | ● | ||
Late | Hyaloscyphaceae (SH0973189.10FU) | 0.707 | 0.006 | ● | ● |
Rozellomycota (SH0910702.10FU) | 0.707 | 0.006 | ● | ||
Ilyonectria mors-panacis (SH1450888.10FU) | 0.707 | 0.007 | ● | ||
Rozellomycota (OTU 1883) | 0.711 | 0.007 | ● | ||
Pezizomycotina (SH0755218.10FU) | 0.808 | 0.008 | ● | ● | |
Trichoderma (OTU 4045) | 0.707 | 0.008 | ● | ||
Hypocreales (OTU 2917) | 0.707 | 0.009 | ● | ||
Leotiomycetes (SH0948543.10FU) | 0.707 | 0.010 | ● | ● |
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Dierickx, G.; Tondeleir, L.; Asselman, P.; Vandekerkhove, K.; Verbeken, A. What Quality Suffices for Nanopore Metabarcoding? Reconsidering Methodology and Ectomycorrhizae in Decaying Fagus sylvatica Bark as Case Study. J. Fungi 2024, 10, 708. https://doi.org/10.3390/jof10100708
Dierickx G, Tondeleir L, Asselman P, Vandekerkhove K, Verbeken A. What Quality Suffices for Nanopore Metabarcoding? Reconsidering Methodology and Ectomycorrhizae in Decaying Fagus sylvatica Bark as Case Study. Journal of Fungi. 2024; 10(10):708. https://doi.org/10.3390/jof10100708
Chicago/Turabian StyleDierickx, Glen, Lowie Tondeleir, Pieter Asselman, Kris Vandekerkhove, and Annemieke Verbeken. 2024. "What Quality Suffices for Nanopore Metabarcoding? Reconsidering Methodology and Ectomycorrhizae in Decaying Fagus sylvatica Bark as Case Study" Journal of Fungi 10, no. 10: 708. https://doi.org/10.3390/jof10100708
APA StyleDierickx, G., Tondeleir, L., Asselman, P., Vandekerkhove, K., & Verbeken, A. (2024). What Quality Suffices for Nanopore Metabarcoding? Reconsidering Methodology and Ectomycorrhizae in Decaying Fagus sylvatica Bark as Case Study. Journal of Fungi, 10(10), 708. https://doi.org/10.3390/jof10100708