1887

Abstract

Phytoplasmas associated with the plant diseases ash yellows (AshY, occurring in ) and lilac witches’-broom (LWB, occurring in ) represent a putative species-level taxon. Phytoplasmal DNA from 19 ash or lilac sources across the known geographic range of AshY (71–113 °W) was examined to determine if AshY and LWB phytoplasmas are a coherent group, if variability exists in both conserved and anonymous DNA, and if variability in 16S rDNA is related to host or geographic origin. The 16S rRNA gene and the 16S-23S spacer were amplified using primer pair P1/P7 and analysed using 15 restriction enzymes. RFLPs were detected in digests obtained with or Taql, for a total of four RFLP profile types. Sequencing of the amplimers from strains AshY1, AshY3, AshY5 and LWB3 (which represent the four 16S rDNA RFLP profile types) revealed only three positions in the 16S rRNA gene and one position in the 16S-23S spacer at which differences occurred; these were single nucleotide substitutions. Sequence homology between any two strains was >99.8%. A portion of a ribosomal protein operon, amplified with primer pair rpF1/R1 from each of the four strains noted above, was analysed with six restriction enzymes, resulting in the detection of two RFLP profiles with . Southern analysis, utilizing two non-specific probes from other phytoplasma groups, revealed three RFLP profile types in anonymous chromosomal DNA of strains representing the four 16S rDNA genotypes. Two strains, AshY3 and LWB3, had unique combinations of characters in the various assays. On the basis of RFLP profiles, the strains from the other plants sampled comprised two groups. The grouping was not clearly related to host or geographic origin. The genome size of strain AshY3 was estimated from PFGE data to be 645 kbp. Phylogenetic analysis of a 1423 bp 16S rDNA sequence from strains AshY1, AshY3, AshY5 and LWB3, together with sequences from 14 other mollicutes archived in GenBank, produced a tree on which the AshY and LWB strains clustered as a discrete group, consistent with previous analyses utilizing only type strain AshY1. Thus, the AshY phytoplasma group is coherent but heterogeneous. The name ‘Candidatus Phytoplasma fraxini’ is proposed for this group.

Loading

Article metrics loading...

/content/journal/ijsem/10.1099/00207713-49-4-1605
1999-10-01
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/ijsem/49/4/ijs-49-4-1605.html?itemId=/content/journal/ijsem/10.1099/00207713-49-4-1605&mimeType=html&fmt=ahah

References

  1. Bove J. M., Garnier M. 1998; Walled and wall-less eubacteria from plants: sieve-tube-restricted plant pathogens. Plant Cell Tissue Organ Cult 52:7–16
    [Google Scholar]
  2. Davis R. E., Sinclair W. A., Lee l.-M., Dally E. L. 1992; Cloned DNA probes specific for detection of a mycoplasmalike organism associated with ash yellows. Mol Plant-Microbe Interact 5:163–169
    [Google Scholar]
  3. Dellaporta S. L., Wood J., Hicks J. B. 1983; A plant DNA minipreparation: Version II. Plant Mol Biol Rep 1:19–21
    [Google Scholar]
  4. Deng S., Hiruki C. 1991; Amplification of 16S rRNA genes from culturable and nonculturable Mollicutes. J Microbiol Methods 14:53–61
    [Google Scholar]
  5. DNASTAR 1997 LASERGENE Biocomputing Software for the Macintosh Madison, WI: DNASTAR;
    [Google Scholar]
  6. Firrao G., Smart C. D., Kirkpatrick B. C. 1996; Physical map of western X-disease phytoplasma chromosome. J Bacteriol 178:3985–3988
    [Google Scholar]
  7. Griffiths H. M., Sinclair W. A., Treshow M., Davis R. E., Lee l.-M., Guo Y., Chen T. A. 1994; Characterization of mycoplasmalike organisms from Fraxinus, Syringa, and associated plants from geographically diverse sites. Phytopathology 84:119–126
    [Google Scholar]
  8. Gundersen D. E., Lee l.-M., Rehner S. A., Davis R. E., Kingsbury D. T. 1994; Phylogeny of mycoplasmalike organisms (phytoplasmas): a basis for their classification. J Bacteriol 176:5244–5254
    [Google Scholar]
  9. Gundersen D. E., Lee l.-M., Schaff D. A., Harrison N. A., Chang C. J., Davis R. E., Kingsbury D. T. 1996; Genomic diversity and differentiation among phytoplasma strains in 16S rRNA groups I (aster yellows and related phytoplasmas) and III (X-disease and related phytoplasmas). Int J Syst Bacteriol 46:64–75
    [Google Scholar]
  10. Hibben C. R., Sinclair W. A., Davis R. E., Alexander J. H. I. 1991; Relatedness of mycoplasmalike organisms associated with ash yellows and lilac witches’-broom. Plant Dis 75:1227–1230
    [Google Scholar]
  11. Kirkpatrick B. C. 1997; Phytoplasmas: the evolution of a molecular taxonomic genus. Nachrbl. Dtsch PflanzenschutzdAS126–130
    [Google Scholar]
  12. Lee l.-M., Davis R. E., Dewitt N. D. 1990; Nonradioactive screening method for isolation of disease-specific probes to diagnose plant diseases caused by mycoplasmalike organisms. Appl Environ Microbiol 56:1471–1475
    [Google Scholar]
  13. Lee l.-M., Davis R. E., Hiruki C. 1991; Genetic interrelatedness among clover proliferation mycoplasmalike organisms (MLOs) and other MLOs investigated by nucleic acid hybridization and restriction fragment length polymorphism analyses. Appl Environ Microbiol 57:3565–3569
    [Google Scholar]
  14. Lee l.-M., Gundersen D. E., Davis R. E., Chiykowski L. N. 1992; Identification and analysis of a genomic strain cluster of mycoplasmalike organisms associated with Canadian peach (eastern) X disease, western X disease, and clover yellow edge. J Bacteriol 174:6694–6698
    [Google Scholar]
  15. Lee l.-M., Davis R. E., Sinclair W. A., Dewitt N. D., Conti M. 1993a; Genetic relatedness of mycoplasmalike organisms detected in Ulmus spp. in the United States and Italy by means of DNA probes and polymerase chain reactions. Phytopathology 83:829–833
    [Google Scholar]
  16. Lee l.-M., Hammond R. W., Davis R. E., Gundersen D. E. 1993b; Universal amplification and analysis of pathogen 16S rDNA for classification and identification of mycoplasmalike organisms. Phytopathology 83:834–842
    [Google Scholar]
  17. Lee l.-M., Gundersen-Rindal D. E., Davis R. E., Bartoszyk I. M. 1998; Revised classification scheme of phytoplasmas based on RFLP analyses of 16S rRNA and ribosomal protein gene sequences. Int J Syst Bacteriol 48:1153–1169
    [Google Scholar]
  18. Li H., Qiu B., Shi C., Jin K., Zhou Q., Huang X. 1997; PCR amplification of 16S rDNA of phytoplasma associated with cherry fasciated disease and RFLP analysis. For Res 10:478–181
    [Google Scholar]
  19. Lim P. O., Sears B. B. 1989; 16S rRNA sequence indicates that plant-pathogenic mycoplasmalike organisms are evolutionarily distinct from animal mycoplasmas. J Bacteriol 171:5901–5906
    [Google Scholar]
  20. Lim P. O., Sears B. B. 1992; Evolutionary relationships of a plant-pathogenic mycoplasmalike organism and Acholeplasma laidlawii deduced from two ribosomal protein gene sequences. J Bacteriol 174:2606–2611
    [Google Scholar]
  21. McCoy R. E., Caudwell A., Chang C. J.16 others 1989 Plant diseases associated with mycoplasmalike organisms. The Mycoplasmas 5545–640 Spiroplasmas, Acholeplasmas, and Mycoplasmas of Plants and Arthropods Edited by Whitcomb R. F., Tully J. G. New York: Academic Press;
    [Google Scholar]
  22. Maddison W. P., Maddison D. R. 1992 MACCLADE’. Analysis of Phylogeny and Character Evolution version 3.03 Sunderland, MA: Sinauer Associates;
    [Google Scholar]
  23. Maurer R., Seemliller E., Sinclair W. A. 1993; Genetic relatedness of mycoplasma-like organisms affecting elm, alder, and ash in Europe and North America. Phytopathology 83:971–976
    [Google Scholar]
  24. Murray R. G. E., Stackebrandt E. 1995; Taxonomic note: implementation of the provisional status Candidates for incompletely described procaryotes. Int J Syst Bacterial 45:186–187
    [Google Scholar]
  25. Nakashima K., Kato S., Iwanami S., Murata N. 1993; DNA probes reveal relatedness of rice yellow dwarf mycoplasmalike organisms (MLOs) and distinguish them from other MLOs. Appl Environ Microbiol 59:1206–1212
    [Google Scholar]
  26. Namba S., Oyaizu H., Kato S., Iwanami S., Tsuchizaki T. 1993; Phylogenetic diversity of phytopathogenic mycoplasmalike organisms. Int J Syst Bacterial 43:461–467
    [Google Scholar]
  27. Neimark H., Carle P. 1995 Mollicute chromosome size determination and characterization of chromosomes from uncultured mollicutes. Molecular and Diagnostic Procedures in Mycoplasmology 1119–131 Edited by Razin S., Tully J. G. San Diego, CA: Academic Press;
    [Google Scholar]
  28. Neimark H., Kirkpatrick B. C. 1993; Isolation and characterization of full-length chromosomes from non-culturable plant-pathogenic Mycop/asTm-hke organisms. Mol Microbiol 7:21–28
    [Google Scholar]
  29. Neimark H. C., Lange C. S. 1990; Pulse-field electrophoresis indicates full-length mycoplasma chromosomes range widely in size. Nucleic Acids Res 18:5443–5448
    [Google Scholar]
  30. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual, 2nd. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  31. Schneider B., Cousins M. T., Klinkong S., Seemuller E. 1995a; Taxonomic relatedness and phylogenetic positions of phytoplasmas associated with diseases of faba bean, sunnhemp, sesame, soybean, and eggplant. Z Pflanzenkr Pflanzensch 102:225–232
    [Google Scholar]
  32. Schneider B., Seemüller E., Smart C. D., Kirkpatrick B. C. 1995b Phylogenetic classification of plant pathogenic mycoplasma-like organisms or phytoplasmas. Molecular and Diagnostic Procedures in Mycoplasmology 1369–380 Edited by Razin S., Tully J. G. San Diego, CA: Academic Press;
    [Google Scholar]
  33. Schneider B., Gibb K. S., Seemüller E. 1997; Sequence and RFLP analysis of the elongation factor Tu gene used in differentiation and classification of phytoplasmas. Microbiology 143:3381–3389
    [Google Scholar]
  34. Seemüller E. 1976; Investigations to demonstrate mycoplasmalike organisms in diseased plants by fluorescence microscopy. Acta Hortic 67:109–112
    [Google Scholar]
  35. Seemüller E., Schneider B., Mäurer R.8 other authors 1994; Phylogenetic classification of phytopathogenic mollicutes by sequence analysis of 16S ribosomal DNA. Int J Syst Bacterial 44:440–446
    [Google Scholar]
  36. Seemüller E., Marcone C, Lauer U., Ragozzino A., Göschl M. 1998; Current status of molecular classification of phytoplasmas. J Plant Pathol 80:3–26
    [Google Scholar]
  37. Sinclair W. A., Griffiths H. M. 1994; Ash yellows and its relationship to dieback and decline of ash. Annu Rev Phytopathol 32:49–60
    [Google Scholar]
  38. Sinclair W. A., Griffiths H. M., Davis R. E. 1996; Ash yellows and lilac witches’-broom: phytoplasmal diseases of concern in forestry and horticulture. Plant Dis 80:468–475
    [Google Scholar]
  39. Smart C. D., Schneider B., Blomquist C. L, Guerra L. J., Harrison N. A., Ahrens U., Lorenz K.-H., Seemüller E., Kirkpatrick B. C. 1996; Phytoplasma-specific PCR primers based on sequences of the 16S-23S rRNA spacer region. Appl Environ Microbiol 62:2988–2993
    [Google Scholar]
  40. Swofford D. L. 1993 paup, version 3.1.1. Champaign, IL: Illinois Natural History Survey;
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/ijsem/10.1099/00207713-49-4-1605
Loading
/content/journal/ijsem/10.1099/00207713-49-4-1605
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error