Investigating unexplained genetic variation and its expression in the arbuscular mycorrhizal fungus Rhizophagus irregularis: A comparison of whole genome and RAD sequencing data

Autoři: Frédéric G. Masclaux aff001;  Tania Wyss aff001;  Marco Pagni aff002;  Pawel Rosikiewicz aff001;  Ian R. Sanders aff001
Působiště autorů: Department of Ecology and Evolution, University of Lausanne, Switzerland aff001;  Vital-IT Group, Swiss Institute of Bioinformatics, Switzerland aff002
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article


Arbuscular mycorrhizal fungi (AMF) are important symbionts of plants. Recently, studies of the AMF Rhizophagus irregularis recorded within-isolate genetic variation that does not completely match the proposed homokaryon or heterokaryon state (where heterokaryons comprise a population of two distinct nucleus genotypes). We re-analysed published data showing that bi-allelic sites (and their frequencies), detected in proposed homo- and heterokaryote R. irregularis isolates, were similar across independent studies using different techniques. This indicated that observed within-fungus genetic variation was not an artefact of sequencing and that such within- fungus genetic variation possibly exists. We then looked to see if bi-allelic transcripts from three R. irregularis isolates matched those observed in the genome as this would give a strong indication of whether bi-allelic sites recorded in the genome were reliable variants. In putative homokaryon isolates, very few bi-allelic transcripts matched those in the genome. In a putative heterokaryon, a large number of bi-allelic transcripts matched those in the genome. Bi-allelic transcripts also occurred in the same frequency in the putative heterokaryon as predicted from allele frequency in the genome. Our results indicate that while within-fungus genome variation in putative homokaryon and heterokaryon AMF was highly similar in 2 independent studies, there was little support that this variation is transcribed in homokaryons. In contrast, within-fungus variation thought to be segregated among two nucleus genotypes in a heterokaryon isolate was indeed transcribed in a way that is proportional to that seen in the genome.

Klíčová slova:

Fungal genetics – Fungal genomics – Fungi – Genetic polymorphism – Genome sequencing – RNA isolation – RNA sequencing – Transcriptome analysis


1. van der Heijden MGA, Martin FM, Selosse M-A, Sanders IR. Mycorrhizal ecology and evolution: the past, the present, and the future. New Phytol. 2015;205:1406–23. doi: 10.1111/nph.13288 25639293.

2. Keymer A, Pimprikar P, Wewer V, Huber C, Brands M, Bucerius SL, et al. Lipid transfer from plants to arbuscular mycorrhiza fungi. eLife. 2017;6:1–33. doi: 10.7554/eLife.29107 28726631.

3. Sanders IR, Croll D. Arbuscular mycorrhiza: the challenge to understand the genetics of the fungal partner. Ann Rev Gen. 2010;44:271–92. doi: 10.1146/annurev-genet-102108-134239 20822441.

4. Ehinger MO, Croll D, Koch AM, Sanders IR. Significant genetic and phenotypic changes arising from clonal growth of a single spore of an arbuscular mycorrhizal fungus over multiple generations. New Phytol. 2012;196:853–61. doi: 10.1111/j.1469-8137.2012.04278.x 22931497.

5. Angelard C, Colard A, Niculita-Hirzel H, Croll D, Sanders IR. Segregation in a mycorrhizal fungus alters rice growth and symbiosis-specific gene transcription. Curr Biol. 2010;20:1216–21. doi: 10.1016/j.cub.2010.05.031 20541408.

6. Sanders IR. Sex, plasticity, and biologically significant variation in one Glomeromycotina species. New Phytol. 2018;220:968–70. doi: 10.1111/nph.15049 29480929

7. Boon E, Zimmerman E, St-Arnaud M, Hijri M. Allelic differences within and among sister spores of the arbuscular mycorrhizal fungus Glomus etunicatum suggest segregation at sporulation. Plos One. 2013;8(12). doi: 10.1371/journal.pone.0083301 24386173.

8. Kuhn G, Hijri M, Sanders IR. Evidence for the evolution of multiple genomes in arbuscular mycorrhizal fungi. Nature. 2001;414:745–8. doi: 10.1038/414745a 11742398.

9. Hijri M, Sanders IR. Low gene copy number shows that arbuscular mycorrhizal fungi inherit genetically different nuclei. Nature. 2005;433:160–3. doi: 10.1038/nature03069 15650740.

10. Pawlowska TE, Taylor JW. Organization of genetic variation in individuals of arbuscular mycorrhizal fungi. Nature. 2004;427:733–7. doi: 10.1038/nature02290 14973485.

11. Boon E, Zimmerman E, Lang BF, Hijri M. Intra-isolate genome variation in arbuscular mycorrhizal fungi persists in the transcriptome. J Evol Biol. 2010;23:1519–27. doi: 10.1111/j.1420-9101.2010.02019.x 20492090.

12. Boon E, Halary S, Bapteste E, Hijri M. Studying genome heterogeneity within the arbuscular mycorrhizal fungal cytoplasm. Genome Biol Evol. 2015;7:505–21. doi: 10.1093/gbe/evv002 25573960.

13. Wyss T, Masclaux FG, Rosikiewicz P, Pagni M, Sanders IR. Population genomics reveals that within-fungus polymorphism is common and maintained in populations of the mycorrhizal fungus Rhizophagus irregularis. ISME J. 2016:1–13. doi: 10.1038/ismej.2016.29 26953600.

14. Lin K, Limpens E, Zhang Z, Ivanov S, Saunders DGO, Mu D, et al. Single nucleus genome sequencing reveals high similarity among nuclei of an endomycorrhizal fungus. PLoS Gen. 2014;10:e1004078. doi: 10.1371/journal.pgen.1004078 24415955.

15. Ropars J, Toro KS, Noel J, Pelin A, Charron P, Farinelli L, et al. Evidence for the sexual origin of heterokaryosis in arbuscular mycorrhizal fungi. Nature Microbiol. 2016;1:16033. doi: 10.1038/nmicrobiol.2016.33 27572831.

16. Tisserant E, Malbreil M, Kuo A, Kohler A, Symeonidi A, Balestrini R, et al. Genome of an arbuscular mycorrhizal fungus provides insight into the oldest plant symbiosis. Proc Natl Acad Sci USA. 2013;110:20117–22. doi: 10.1073/pnas.1313452110 24277808.

17. Savary R, Masclaux FG, Wyss T, Droh G, Cruz Corella J, Machado AP, et al. A population genomics approach shows widespread geographical distribution of cryptic genomic forms of the symbiotic fungus Rhizophagus irregularis. ISME J. 2018;12:17–30. doi: 10.1038/ismej.2017.153 29027999.

18. Maeda T, Kobayashi Y, Kameoka H, Okuma N, Takeda N, Yamaguchi K, et al. Evidence of non-tandemly repeated rDNAs and their intragenomic heterogeneity in Rhizophagus irregularis. Commun Biol. 2018;1:87. doi: 10.1038/s42003-018-0094-7 30271968.

19. Doyle JJ, Flagel LE, Paterson AH, Rapp RA, Soltis DE, Soltis PS, et al. Evolutionary genetics of genome merger and doubling in plants. Ann Rev Gen. 2008;42:443–61. doi: 10.1146/annurev.genet.42.110807.091524 18983261.

20. Bécard G, Fortin JA. Early events of vesicular-arbuscular mycorrhiza formation on Ri T-DNA transformed roots. New Phytol. 1988;108:211–8.

21. Rosikiewicz P, Bonvin J, Sanders IR. Cost-efficient production of in vitro Rhizophagus irregularis. Mycorrhiza. 2017;27:477–86. doi: 10.1007/s00572-017-0763-2 28210812.

22. Schmieder R, Lim YW, Rohwer F, Edwards R. TagCleaner: Identification and removal of tag sequences from genomic and metagenomic datasets. BMC Bioinformatics. 2010;11:341. doi: 10.1186/1471-2105-11-341 20573248.

23. Schmieder R, Edwards R. Quality control and preprocessing of metagenomic datasets. Bioinformatics. 2011;27:863–4. doi: 10.1093/bioinformatics/btr026 21278185.

24. Pearson WR, Lipman DJ. Improved tools for biological sequence comparison. Proc Natl Acad Sci USA. 1988;85:2444–8. doi: 10.1073/pnas.85.8.2444 3162770.

25. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, et al. The sequence alignment/map format and SAMtools. Bioinformatics. 2009;25:2078–9. doi: 10.1093/bioinformatics/btp352 19505943.

26. Garrison E, Marth G. Haplotype-based variant detection from short-read sequencing. 2012:1–9. Preprint. Available from: arXiv:1207.3907 [q-bio.GN]. Cited 1 July 2019.

27. Conway JR, Lex A, Gehlenborg N. UpSetR: an R package for the visualization of intersecting sets and their properties. Bioinformatics. 2017;33:2938–40. doi: 10.1093/bioinformatics/btx364 28645171.

28. Sacomoto GAT, Kielbassa J, Chikhi R, Uricaru R, Antoniou P, Sagot MF, et al. KisSplice: De-novo calling alternative splicing events from RNA-seq data. BMC Bioinformatics. 2012;13:S5. doi: 10.1186/1471-2105-13-S6-S5 22537044.

29. Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, et al. Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nature Biotechnology. 2011;29:644–52. doi: 10.1038/nbt.1883 21572440.

30. Chen ECH, Morin E, Beaudet D, Noel J, Yildirir G, Ndikumana S, et al. High intraspecific genome diversity in the model arbuscular mycorrhizal symbiont Rhizophagus irregularis. New Phytol. 2018;220(4):1161–71. doi: 10.1111/nph.14989 29355972

31. Chen EC, Mathieu S, Hoffrichter A, Sedzielewska-Toro K, Peart M, Pelin A, et al. Single nucleus sequencing reveals evidence of inter-nucleus recombination in arbuscular mycorrhizal fungi. eLife. 2018;7:1–17. doi: 10.7554/eLife.39813 30516133.

32. Masclaux FG, Wyss T, Mateus-Gonzalez ID, Aletti C, Sanders IR. Variation in allele frequencies at the bg112 locus reveals unequal inheritance of nuclei in a dikaryotic isolate of the fungus Rhizophagus irregularis. Mycorrhiza. 2018;28:369–77. doi: 10.1007/s00572-018-0834-z 29675619.

33. Gehmann T, Pelkmans J, Ohm R, Vos A, Sonnenberg A, Baars J, et al. Nucleus-specific expression in the multinuclear mushroom-forming fungus Agaricus bisporus reveals different nuclear regulatory programs. Proc Natl Acad Sci USA. 2018;115:4429–34. doi: 10.1073/pnas.1721381115 29643074.

Článek vyšel v časopise


2019 Číslo 12
Nejčtenější tento týden