Exploring Possible Variation among Iranian Erwinia amylovora Strains Using Multilocus Typing and Tandem Repeat Analysis

Authors
Department of Crop Protection, Faculty of Agriculture, Ferdowsi University of Mashhad, P. O. Box: 91775-1163, Mashhad, Islamic Republic of Iran.
Abstract
In the current study, molecular typing of 50 Erwinia amylovora strains related to different regions in Iran was evaluated using multi-locus sequence analysis and variable number of tandem repeats. In the first assay, phylogenetic tree based on partial sequences of recombinase A, sigma factor S and a heat shock protein GroEL showed significant identity in studied gene sequences. A single nucleotide variation in groEL was determined in IrGh59 strain related to Crataegus spp. from Ghazvin Province. In VNTR analysis, the same fingerprinting profile similar to E. amylovora reference strain ATCC49946 was yielded for tested strains except NBQ1 and MQ1 which may reflect a unique contaminating source for this disease in Iran. In addition, the honey-bee movements with respect to blossom season probably have a considerable role in fire blight unique dispersal in our area. The NBQ1 and MQ1 strains generated different VNTR profiles, isolated from cultivars NeishabourandEsfehan of Cydonia oblonga plant, respectively. No definite assessment can be expressed in this case. However, possible entry of other infection mass from neighboring countries should be determined. Overall, VNTR profile analyses are recommended as a tool to evaluate genetic differences in E. amylovora populations. In addition, employing more strains from different known sources could be assistance to achieve more accurate results about E. amylovora genetic variation and also fire blight distribution patterns.

Keywords


1. Bereswill, S., Pahl, A., Bellemann, P., Zeller, W. and Geider, K. 1992. Sensitive and Species-Specific Detection of Erwinia amylovora by PCR-Analysis. Appl. Environ. Microbiol., 58: 3522–3526.
2. Bühlmann, A., Dreo, T., Rezzonico, F., Pothier, J. F., Smits, T. H. M., Ravnikar, M., Frey, J. E. and Duffy, B. 2013. Phylogeography and Population Structure of the Biologically Invasive Phytopathogen Erwinia amylovora Inferred Using Minisatellites. Environ. Microbiol., 16: 2112-25.
3. Donat, V., Biosca, E. G., Penalver, J. and Lopez, M. M. 2007. Exploring Diversity among Spanish Strains of Erwinia amylovora and Possible Infection Sources. J. Appl. Microbiol., 103: 1639-1649.
4. Dreo, T., Ravnikar, M., Frey, J. E., Smits, T.H.M., Duffy, B. 2011. Silico Analysis of Variable Number of Tandem Repeats in Erwinia Amylovora Genomes. Acta Hortic.896: 115-118.
5. Gehring, I. and Geider, K. 2012. Differentiation of Erwinia amylovora and Erwinia pyrifoliae Strains with Single Nucleotide Polymorphisms and by Synthesis of dihydrophenylalanine. Curr. Microbiol., 65: 73–84.
6. Geider, K., Auling, G., Jakovljevic, V. and Volksch, B. 2009. A Polyphasic Approach Assigns the Pathogenic Erwinia Strains from Diseased Pear Trees in Japan to Erwinia pyrifoliae. Lett. Appl. Microbiol., 48: 324-330.
7. Hannou, N., Llop, P., Faure, D., Lopez, M. M. and Moumni, M. 2013. Characterization of Erwinia amylovora Strains from Middle Atlas Mountains in Morocco by PCR Based on Tandem Repeat Sequences. Eur. J. Plant Pathol., 136: 665-674.
8. Jock, S., Donat, V., Lopez, M. M., Bazzi, C., Geider, K. 2002. Following Spread of Fire Blight in Western, Central and Southern Europe by Molecular Differentiation of Erwinia amylovora Strains with PFGE Analysis. Environ. Microbiol., 4: 106–114.
9. Llop, P., Barbe, S. and Lopez, M. M. 2012. Functions and Origin of Plasmids in Erwinia Species that Are Pathogenic to or Epiphytically Associated with Pome Fruit. Tree., 26: 31-46.
10. Maiden, M. C., Bygraves, J. A., Feil, E., Morelli, G., Russell, J. E. and Urwin, R. Zhang, Q., Zhou, J., Zurth, K., Caugant, D. A., Feavers, I. M., Achtman, M., Spratt, B.G.. 1998. Multilocus Sequence Typing: A Portable Approach to the Identification of Clones within Populations of Pathogenic Microorganisms. Proc. Natl. Acad. Sci. USA., 95: 3140–3145.
11. Mansfield, J., Genin, S., Magori, S., Citovsky, V. ,Sriariyanum, M. and Ronald, P. Dow, M., Verdier, V., Beer, S. V., Machado, M. A., Toth, I., Salmond, G., Foster, G. D. 2012. Top 10 Plant Pathogenic Bacteria in Molecular Plant Pathology. Mol. Plant Pathol. 13: 614–629.
12. McGhee, G. C., Guasco, J., Bellomo, L. M., Blumer-Schuette, S. E., Shane, W. W., Irish-Brown, A. and Sundin, G. W. 2011. Genetic Analysis of Streptomycin-Resistant (Sm-R) Strains of Erwinia amylovora Suggests that Dissemination of Two Genotypes Is Responsible for the Current Distribution of Sm-R E. amylovorain Michigan. Phytopathol., 101: 182–191.
13. McGhee, G. C., Schnabel, E. L., Maxson-Stein, K., Jones, B., Stromberg, V. K., Lacy, G. H. and Jones, A. L. 2002. Relatedness of Chromosomal and Plasmid DNAs of Erwinia pyrifoliae and Erwinia amylovora. Appl. Environ. Microbiol., 68: 6182-6192.
14. McManus, P. S. and Jones, A. L. 1995. Genetic Fingerprinting of Erwinia amylovora Strains Isolated from Tree-Fruit Crops and Rubus spp. Phytopathol., 85: 1547–1553.
15. Mohammadi, M. 2010. Enhanced Colonization and Pathogenicity of Erwinia amylovora Strains Transformed with the Near-Ubiquitous pEA29 Plasmid on Pear and Apple. Plant Pathol,. 59: 252–261.
16. Momol, M. T., Momol, E. A., Lamboy, W. F., Norelli, J. L., Beer, S. V. and Aldwinckle, H. S. 1997. Characterization of Erwinia amylovora Strains Using Random Amplified Polymorphic DNA Fragments (RAPDs). J. Appl. Microbiol., 82: 339–398.
17. Palacio-Bielsa, A., Rosello, M., Llop, P., Lopez, M. M. 2012. Erwinia spp. from Pome Fruit Trees: Similarities and Differences among Pathogenic and Non-Pathogenic Species. Tree., 26: 13–29.
18. Powney, R., Smits, T. H. M., Sawbridge, T., Frey, B., Blom, J., Frey, J. E., Plummer, K. M., Beer, S. V., Luck, J., Duffy, B. and Rodoni, B. 2011. Genome Sequence of an Erwinia amylovora Strain with Pathogenicity Restricted to Rubus Plants. J. Bacteriol., 193: 785-786.
19. Puławska, J. and Sobiczewski, P. 2012. Phenotypic and Genetic Diversity of Erwinia amylovora: The Causal Agent of Fire Blight. Trees, 26: 3–12.
20. Schaad, N. W., Jones, J. B. and Chun, W. 2001. Laboratory Guide for Identification of Plant Pathogenic Bacteria. APS Press, St. Paul, MN, USA, 158 PP.
21. Sebaihia, M., Bocsanczy, A. M., Biehl, B.S., Quail, M. A., Pema, N. T., Glasner, J. D., DeClerck, G. A., Cartinjour, S., Schneider, D. J., Bentley, S. D., Parkhill, J. and Beer, S. V. 2010.Complete Genome of the Plant Pathogen Erwinia amylovora Strain ATCC 49946. J. Bacteriol., 192: 2020-2021.
22. Shrestha, R., Lee, S. H., Kim, J. E., Wilson, C., Choi, S. G, Park, D. H., Wang, M. H., Hur, J. H., Lim, C. K. 2007. Diversity and Detection of Korean Erwinia pyrifoliae Strains as Determined by Plasmid Profiling, Phylogenetic Analysis and PCR. Plant Pathol., 56:1023–1031.
23. Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M. and Kumar, S. 2011. MEGA5: Molecular Evolutionary Genetics Analysis Using Maximum Like-Lihood, Evolutionary Distance, and Maximum Parsimony Methods. Mol. Biol. Evol., 28: 2731-2739.
24. Taghdareh, Gh., Baghaee-Ravari, S., Moslemkhani, C. and Mahdikhani-Moghaddam, E. 2014. Evaluation of Repeat Sequences on Plasmid pEA29 of Erwinia amylovora from Iran. Eur. J. Plant Pathol., 140: 735–744.
25. Taylor, R. K., Guilford, P. J., Clark, R. G., Hale, C. N. and Forster, R. L. S. 2001. Detection of Erwinia amylovora in Plant Material Using Novel Polymerase Chain Reaction (PCR) Orimers. New Zealand J. Crop Hort. Sci., 29: 35–43.
26. Thompson, J. D., Gibson, T. J., Plewniak, F. and Higgins, D. G. 1997. The CLUSTALX Windows Interface: Flexible Strategies for Multiple Sequence Alignment Aided by Quality Analysis Tools. Nucl. Acid. Res., 24: 4876–4882.
27. Van Belkum, A. 1999. The Role of Short Sequence Repeats in Epidemiologic Typing. Curr. Opin. Microbiol., 2: 306–311.
28. Waleron, M., Waleron, K., Geider, K. and Lojkowska, E. 2008. Application of RFLP Analysis of recA, gyrA and rpoS Gene Fragments for Rapid Differentiation of Erwinia amylovora from Erwinia Strains Isolated in Korea and Japan. Eur. J. Plant Pathol., 121: 161–172.
29. Waleron, M., Waleron, K. and Łojkowska, E. 2002b. Genotypic Characterization of the Erwinia Genus by PCR-RFLP Snalysis of rpoS Gene. Plant Protec. Sci., 38: 288–290.
30. Waleron, M., Waleron, K., Podhajska, A. J. and Lojkowska, E. 2002a. Genotyping of Bacteria Belonging to the Former Erwinia Genus by PCR–RFLP Analysis of a recA Gene Fragment. Microbiology, 148: 583–595.