Skip to main content

Advertisement

SpringerLink
Log in

Identification and characterization of polymorphic minisatellites in the phytopathogenic ascomycete Leptosphaeria maculans

  • Research Article
  • Published:
Current Genetics Aims and scope Submit manuscript

Abstract

Leptosphaeria maculans causes phoma stem canker, the most serious disease of oilseed rape world-wide. Sexual recombination is important in the pathogen life cycle and increases the risk of plant resistance genes being overcome rapidly. Thus, there is a need to develop easy-to-use molecular markers suitable for large-scale population genetic studies. The minisatellite MinLm1, showing six alleles in natural populations, has previously been used as a marker to survey populations. Here, we report the characterization of five new minisatellites (MinLm2–MinLm6), of which four were identified by a systematic search for tandemly repeated polymorphic regions in BAC-end sequencing data from L. maculans. Of 782 BAC-end sequences analysed, 43 possessed putative minisatellite-type repeats and four of these (MinLm3–MinLm6) displayed both consistent PCR amplification and size polymorphism in a collection of L. maculans isolates of diverse origins. Cloning and sequencing of each allele confirmed that polymorphism was due to variation in the repeat number of a core motif ranging from 11 bp (MinLm3) to 51 bp (MinLm4). The number of alleles found for each minisatellite ranged from three (MinLm4) to nine (MinLm2), with eight, five and six for MinLm3, MinLm5 and MinLm6, respectively. MinLm2–MinLm6 are all single locus markers specific to L. maculans and share some common features, such as conservation of core motifs and incomplete direct repeats in the flanking regions. To our knowledge, L. maculans is the first fungal species for which six polymorphic single locus minisatellite markers have been reported.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Andersen TH, NilssonTillgren T (1997) A fungal minisatellite. Nature 386:771

    Article  CAS  PubMed  Google Scholar 

  • Armour JAL, Jeffreys AJ (1992) Recent advances in minisatellite biology. FEBS Lett 307:113–115

    Article  CAS  PubMed  Google Scholar 

  • Attard A, Gourgues M, Gout L, Schmit J, Roux J, Narcy JP, Balesdent MH, Rouxel T (2001) Molecular characterisation and polymorphism of MinLm1, a minisatellite from the phytopathogenic ascomycete Leptosphaeria maculans. Curr Genet 40:54–64

    Article  CAS  PubMed  Google Scholar 

  • Attard A, Gout L, Gourgues M, Kuhn ML, Schmit J, Laroche S, Ansan-Melayah D, Billault A, Cattolico L, Balesdent MH, Rouxel T (2002) Analysis of molecular markers genetically linked to the Leptosphaeria maculans avirulence gene AvrLm1 in field populations indicates a highly conserved event leading to virulence on Rlm1 genotypes. Mol Plant-Microbe Interact 15:672–682

    CAS  PubMed  Google Scholar 

  • Attard A, Gout L, Ross S, Parlange F, Cattolico L, Balesdent M-H, Rouxel T (2004) Truncated and RIP-degenerated copies of the LTR retrotransposon Pholy are clustered in a pericentromeric region of the Leptosphaeria maculans genome. Fungal Genet Biol (in press)

  • Balesdent MH, Attard A, Ansan-Melayah D, Delourme R, Renard M, Rouxel T (2001) Genetic control and host range of avirulence toward Brassica napus cultivars Quinta and Jet Neuf in Leptosphaeria maculans. Phytopathology 91:70–76

    Google Scholar 

  • Balesdent MH, Attard A, Kuhn ML, Rouxel T (2002) New avirulence genes in the phytopathogenic fungus Leptosphaeria maculans. Phytopathology 92:1122–1133

    CAS  Google Scholar 

  • Benson G (1999) Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res 27:573–580

    Article  CAS  PubMed  Google Scholar 

  • Bois P, Jeffreys AJ (1999) Minisatellite instability and germline mutation. Cell Mol Life Sci 55:1636–1648

    Article  CAS  PubMed  Google Scholar 

  • Cambareri EB, Aisner R, Carbon J (1998) Structure of the chromosome VII centromere region in Neurospora crassa: degenerate transposons and simple repeats. Mol Cell Biol 18:5465–5477

    CAS  PubMed  Google Scholar 

  • Coates BS, Hellmich RL, Lewis LC (2002) Allelic variation of a Beauveria bassiana (Ascomycota: Hypocreales) minisatellite is independent of host range and geographic origin. Genome 45:125–132

    Article  CAS  PubMed  Google Scholar 

  • Gall C, Balesdent MH, Robin P, Rouxel T (1994) Tetrad analysis of acid phosphatase, soluble protein patterns, and mating type in Leptosphaeria maculans. Phytopathology 84:1299–1305

    CAS  Google Scholar 

  • Giraud T, Fortini D, Levis C, Brygoo Y (1998) The minisatellite MSB1, in the fungus Botrytis cinerea, probably mutates by slippage. Mol Biol Evol 15:1524–1531

    Google Scholar 

  • Goodwin PH, Annis SL (1991) Rapid identification of genetic variation and pathotype of Leptosphaeria maculans by random amplified polymorphic DNA assay. Appl Environ Microbiol 57:2482–2486

    CAS  PubMed  Google Scholar 

  • Haber JE, Louis EJ (1998) Minisatellite origins in yeast and humans. Genomics 48:132–135

    Article  CAS  PubMed  Google Scholar 

  • Hamann A, Osiewacz HD (1998) Genome analysis of filamentous fungi: identification and characterization of an unusual GT-rich minisatellite in the ascomycete Podospora anserina. Curr Genet 34:88–92

    Article  CAS  PubMed  Google Scholar 

  • Hayden HL, Wilson LM, Cozijnsen AJ, Howlett BJ (2004) Characterisation and cross-species amplification of microsatellite loci in the plant pathogenic fungus Leptosphaeria maculans. Mol Ecol Notes 4:480–481

    Article  CAS  Google Scholar 

  • Horowitz H, Haber JE (1984) Subtelomeric regions of yeast chromosomes contain a 36 base-pair tandemly repeated sequence. Nucleic Acids Res 12:7105–7121

    CAS  PubMed  Google Scholar 

  • Idnurm A, Howlett BJ (2003) Analysis of loss of pathogenicity mutants reveals that repeat-induced point mutations can occur in the Dothideomycete Leptosphaeria maculans. Fungal Genet Biol 39:31–37

    Article  CAS  PubMed  Google Scholar 

  • Jedryczka M, Rouxel T, Balesdent MH (1999) Rep-PCR based genomic fingerprinting of isolates of Leptosphaeria maculans from Poland. Eur J Plant Pathol 105:813–823

    Article  CAS  Google Scholar 

  • Jeffreys AJ, Wilson V, Thein SL (1985) Hypervariable minisatellite regions in human DNA. Nature 314:67–73

    CAS  PubMed  Google Scholar 

  • Kurenova EV, Mason JM (1997) Telomere functions. a review. Biochem Mosc 62:1242–1253

    CAS  Google Scholar 

  • Le Flèche P, Hauck Y, Onteniente L, Prieur A, Denoeud F, Ramisse V, Sylvestre P, Benson G, Ramisse F, Vergnaud G (2001) A tandem repeats database for bacterial genomes: application to the genotyping of Yersinia petis and Bacillus anthracis. BMC Microbiol 1:2

    Article  PubMed  Google Scholar 

  • Leclair S, Ansan-Melayah D, Rouxel T, Balesdent MH (1996) Meiotic behaviour of the minichromosome in the phytopathogenic ascomycete Leptosphaeria maculans. Curr Genet 30:541–548

    Article  CAS  PubMed  Google Scholar 

  • Louis EJ, Naumova ES, Lee A, Naumov G, Haber JE (1994) The chromosome end in yeast—its mosaic nature and influence on recombinational dynamics. Genetics 136:789–802

    CAS  PubMed  Google Scholar 

  • Mannazzu I, Simonetti E, Marinangeli P, Guerra E, Budroni M, Thangavelu M, Clementi F (2002) SED1 gene length and sequence polymorphisms in feral strains of Saccharomyces cerevisiae. Appl Environ Microbiol 68:5437–5444

    CAS  PubMed  Google Scholar 

  • Margolin BS, Garrett-Engele PW, Stevens JN, Fritz DY, Garrett-Engele C, Metzenberg RL, Selker EU (1998) A methylated Neurospora 5S rRNA pseudogene contains a transposable element inactivated by repeat-induced point mutation. Genetics 149:1787–1797

    CAS  PubMed  Google Scholar 

  • Marinangeli P, Angelozzi D, Ciani M, Clementi F, Mannazzu I (2004) Minisatellites in Saccharomyces cerevisiae genes encoding cell wall proteins: a new way towards wine strain characterisation. FEMS Yeast Res 4:427–435

    Article  CAS  PubMed  Google Scholar 

  • Mendes-Pereira E, Balesdent M-H, Brun H, Rouxel T (2003) Molecular phylogeny of the Leptosphaeria maculans–L. biglobosa species complex. Mycol Res 107:1287–1304

    Article  CAS  PubMed  Google Scholar 

  • Nicholas KB, Nicholas HB (1997) Genedoc: a tool for editing and annotating multiple sequence alignments, ver. 2.6.002.

  • Onteniente L, Brisse S, Tassios PT, Vergnaud G (2003) Evaluation of the polymorphisms associated with tandem repeats for Pseudomonas aeruginosa strain typing. J Clin Microbiol 41:4991–4997

    Google Scholar 

  • Parker PG, Snow AA, Schug MD, Booton GC, Fuerst PA (1998) What molecules can tell us about populations: choosing and using a molecular marker. Ecology 79:361–382

    Google Scholar 

  • Pinochet X, Mestries E, Penaud A, Delourme R, Chevre M, Renard M, Brun H, Bousset L, Balesdent M-H, Rouxel T, Aubertot JN (2003) Towards a durable management of genetic resistances to Leptosphaeria maculans. Ocl-Ol Corps Gras Li 10:208–211

    Google Scholar 

  • Plummer KM, Howlett BJ (1993) Major chromosomal length polymorphisms are evident after meiosis in the phytopathogenic fungus Leptosphaeria maculans. Curr Genet 24:107–113

    Article  CAS  PubMed  Google Scholar 

  • Pongam P, Osborn TC, Williams PH (1999) Assessment of genetic variation among Leptosphaeria maculans isolates using pathogenicity data and AFLP analysis. Plant Dis 83:149–154

    Google Scholar 

  • Pourcel C, Vidgop Y, Ramisse F, Vergnaud G, Tram C (2003) Characterization of a tandem repeat polymorphism in Legionella pneumophila and its use for genotyping. J Clin Microbiol 41:1819–1826

    Google Scholar 

  • Pouwels D, Simons G (2003) Fingerprinting microorganisms. Food Technol 57:36–40

    CAS  Google Scholar 

  • Purwantara A, Barrins JM, Cozijnsen AJ, Ades PK, Howlett BJ (2000) Genetic diversity of isolates of the Leptosphaeria maculans species complex from Australia, Europe and North America using amplified fragment length polymorphism analysis. Mycol Res 104:772–781

    Article  CAS  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning—a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.

    Google Scholar 

  • Sippell DW, Hall R (1995) Glucose phosphate isomerase polymorphisms distinguish weakly virulent from highly virulent-strains of Leptosphaeria maculans. Can J Plant Pathol 17:1–6

    CAS  Google Scholar 

  • Taylor JL, Borgmann IE (1994) An unusual repetitive element from highly virulent isolates of Leptosphaeria maculans and evidence of its transfer to a weakly virulent isolate. Mol Plant-Microbe Interact 7:181–188

    CAS  PubMed  Google Scholar 

  • Taylor JS, Breden F (2000) Slipped-strand mispairing at noncontiguous repeats in Poecilia reticulata: a model for minisatellite birth. Genetics 155:1313–1320

    CAS  PubMed  Google Scholar 

  • Vergnaud G, Denoeud F (2000) Minisatellites: mutability and genome architecture. Genome Res 10:899–907

    Article  CAS  PubMed  Google Scholar 

  • Volke B (1999) Leptosphaeria maculans, der Erreger der Wurzelhals- und Stengelfäule an Raps: Verbreitung verschiedener Pathogenitätsgruppen in Europa, Quantifizierung des Befalls und Schadwirkung im Freiland. Georg-August-Universität, Göttingen, pp 1–116

  • Weising K, Nybom H, Wolff K, Meyer W (1995) DNA fingerprinting in plants and fungi. CRC, Boca Raton

    Google Scholar 

  • West JS, Kharbanda PD, Barbetti MJ, Fitt BDL (2001) Epidemiology and management of Leptosphaeria maculans (phoma stem canker) on oilseed rape in Australia, Canada and Europe. Plant Pathol 50:10–27

    Article  Google Scholar 

  • West JS, Balesdent M-H, Rouxel T, Narcy JP, Huang Y-J, Roux J, Steed JM, Fitt BDL, Schmit J (2002) Colonization of winter oilseed rape tissues by A/Tox+ and B/Tox0 Leptosphaeria maculans (phoma stem canker) in France and England. Plant Pathol 51:311–321

    Article  Google Scholar 

  • Wöstemeyer J, Kreibich A (2002) Repetitive DNA elements in fungi (Mycota): impact on genomic architecture and evolution. Curr Genet 41:189–198

    Google Scholar 

Download references

Acknowledgements

Financial support for this project is gratefully acknowledged. M.E. was funded by the European Union (Marie Curie Fellowship HPMT-CT-2001-00395), the Biotechnology and Biological Sciences Research Council and DuPont. M.J. received funds from the State Committee for Scientific research (KBN), Poland, and the Polish–French integrated research action Polonium 2002. All providers of isolates and support staff at PMDV INRA Versailles are thanked for their assistance and thanks are due to the anonymous reviewers for suggesting further interesting elements to discuss.

Author information

Authors and Affiliations

  1. Rothamsted Research, Harpenden, AL5 2JQ, UK

    Maria Eckert & Bruce Fitt

  2. INRA, PMDV, Route de St. Cyr, 78026, Versailles, France

    Lilian Gout, Thierry Rouxel, Françoise Blaise & Marie-Hélène Balesdent

  3. Institute of Plant Genetics, Polish Academy of Sciences, 60-479, Poznan, Poland

    Malgorzata Jedryczka

Authors
  1. Maria Eckert
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Lilian Gout
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Thierry Rouxel
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Françoise Blaise
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Malgorzata Jedryczka
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Bruce Fitt
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Marie-Hélène Balesdent
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Marie-Hélène Balesdent.

Additional information

Communicated by J. Heitman

Electronic Supplementary Material

Rights and permissions

Reprints and permissions

About this article

Cite this article

Eckert, M., Gout, L., Rouxel, T. et al. Identification and characterization of polymorphic minisatellites in the phytopathogenic ascomycete Leptosphaeria maculans. Curr Genet 47, 37–48 (2005). https://doi.org/10.1007/s00294-004-0539-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00294-004-0539-z

Keywords

Search

Navigation