Mikrobiol. Z. 2017; 79(3):44-55. Ukrainian.
doi: https://doi.org/10.15407/microbiolj79.03.044
Phylogenetic Analysis of Pathogenic Bacteria Belongs to the Erwinia Genus –
Agent of Apple’s Diseases in Ukraine
Dankevich L.A.
Zabolotny Institute of Microbiology and Virology, NAS of Ukraine
154 Akad. Zabolotny Str., Kyiv, 03143, Ukraine
Aim. To determine the similarity of 16S pPHK gene nucleotide sequences of 10 isolated Erwinia sp. and 9 collection «Erwinia horticola» strains with typical representatives of the genus Erwinia for their correct identifcation at the species level. Methods. Objects − 10 isolated from diseased tissue of apple Erwinia sp. and 9 collection «Erwinia horticola» strains. DNA copy of the gene 16S pPHK amplifed using universal primers pA and pH. Building a reverse compliment, editing, alignment and matching sequences was performed using Multalin and Sequence Manipulation Suite software. The nucleotide similar sequences of 16S rRNA gene, that placed and for deposited in GenBank, searching was performed using the BLASTN program. The phylogenetic trees constructing was performed using MEGA 5 program. Results. Nucleotide sequence analysis of 16S rRNA gene found a high level of homology (99 - 98 %) of the test strains with typical strains of E. amylovora, E. pyrifoliae, E. piriforinigrans, E. uzenensis, E. billingiae, E. persicina, E. aphidicola species. Conclusions. Considering examined previously the complex of phenotypic properties and determined homology of 16S pPHK gene nucleotide sequences can be argued that most likely similarity have isolated Erwinia sp. and collection «Erwinia horticola» strains with typical E. amylovora and E. pyrifoliae strains. Accoding to the literature, these species are closely related and poorly differentiated based on the homology of the 16S pPHK gene nucleotide sequences. Probably for the final identifcation at the species level the strains, studied by us, and belongs to the Erwinia genus it is necessary to involvement other genotypic properties.
Key words: Erwinia sp., «Erwinia horticola», Erwinia amylovora, the nucleotide sequence of the gene 16S pPHK, phylogenetic analysis.
Full text (PDF, in Ukrainian)
- Ashmawy NA, Zaghloul TI, El-Sabagh AM. Isolation and Molecular characterization of the fre blight pathogen, Erwinia amylovora, isolation from apple and pear orchards in Egypt. Plant Pathology Journal. 2015; 14(3):142-147. https://doi.org/10.3923/ppj.2015.142.147
- Birnboim HC. A rapid alkaline extraction method for isolation of plasmid DNA. Methods Enzymol. 1983; 100(1) 243-255. https://doi.org/10.1016/0076-6879(83)00059-2
- Brenner DJ, Krieg NR, Staley JT, Garrity GM. Bergey’s Manual of Systematic Bacteriology. Vol. 1. 2nd ed. New York; USA: Springer Science+ Business Media; 2005.
- Dankevich LA, Votselko SK, Scherbina TM, Patyka VP. [Phenotypic heterogynicity phytopathogenic bacteria belongs to Erwinia genus – agent of apples bacterial diseases in Ukraine]. Mikrobiol Z. 2016; 78(5) 30-41. Ukrainian.
- Edwars U, Rogal T, Blocker H. et all. Isolation and direct complete nucleotide determination of entire genes. Characterization of gene coding for 16S ribocomal RNA. Nucl. Asids Res. 1989; 17(19):7843-7853. https://doi.org/10.1093/nar/17.19.7843
- Gvozdyak RI, Pasichnyk LA, Yakovleva LM, Moroz SM, Lytvynchuk OO, Zhytkevych NV, Hodos SF, Butsenko LN, L Dankevich LA, Hrynnyk IV, Patyka VP. Phytopathogenic bacteria. Bacterial diseases of plants. Ed. V.P Patyka. Kyiv: “ Interservice”; 2011. Ukrainian.
- Hauben L, Moore ER, Vauterin L, Steenacker M, Mergaert J, Verdonck L, Swing J. Phylogenetic position of phytopathogens within Enterobacteriaceae. Systematic Applied Micribiology. 1998; 21(3):384-397. https://doi.org/10.1016/S0723-2020(98)80048-9
- Kim WS, Gardan L, Rhim SL, Geider K. Erwinia pyrifoliae sp. nov., a novel pathogen that affects Asian pear trees (Pyrus pyrifolia Nakai). Iternational Journal of Systematic Bacteriology. 1999; 49(2):899-906. https://doi.org/10.1099/00207713-49-2-899
- Kudina IV, Lagonenko AL, Evtushenkov AN. [Characterisation of Belarusian isolates of phytopathogenic bacteria Erwinia amylovora]. Treatise of the BSU. 2008; 3:26-34. Russian.
- List of prokaryotic names with standing in nomenclature. www.bacterio.net/erwinia.html
- Lopes MM, Rosello M, Liop P, Ferrer S, Christen R, Gardan L. Erwinia piriforinigrans sp.nov., a novel pathogen that causes necrosis of pear blossoms. Iternational Journal of Systematic Bacteriology. 2011; 61:561-567. https://doi.org/10.1099/ijs.0.020479-0
- Marchuk D, Drumm M, Saulino AS et all. Construction of T-vectors, a rapid and general system for direct cloning of unmodifed PCR product. Nucl. Acid Res. 1990; 19(5):1154-1155. https://doi.org/10.1093/nar/19.5.1154
- Matsuura T, Mizuno A, Tsukamoto T, Shimizu Y, Saito N, Sato S, Kikuchi S, Uzuki T, Azegami K, Sawada H. Erwinia uzenensis sp. nov., a novel pathogen that affects European pear trees (Pyrus communis L.). Iternational Journal of Systematic Bacteriology. 2012; 62(8):1799-1803. https://doi.org/10.1099/ijs.0.032011-0
- McGhee GC, Schnabel EL, Maxson-Stein K, Jones B, Stromberg VK, Lacy GH, Jones AL. Relatedness of chromosomal and plasmid DNAs of Erwinia pyrifoliae and Erwinia amylovora. Applied and environmental microbiology. 2002; 68(12):6182-6192. https://doi.org/10.1128/AEM.68.12.6182-6192.2002
- Mergaert J, Hauben L, C Cnockaert M, Swings J. Reclassifcation of non-pigmented Erwinia herbicola strains from trees as Erwinia billingae sp. nov. Iternational Journal of Systematic Bacteriology. 1999; 49:377-383. https://doi.org/10.1099/00207713-49-2-377
- Pique N, Minana-Galbis D., Merino S., Tomas J. Virulence factor of Erwinia amylovora: a review. Int. J. Mol. Sci. 2015; 16(6):12836-12854.
- Sambrook J, Fritch EF, Maniatis T. Molecular Cloning: a laboratory manual. N.Y.: Cold Spring Harbor Laboratory Press; 1989.
- Soon-Wo Kwong, Seung-Joo Go, Hee-Wan Kang, Jin-Chang Ryu, Jin-Ki Jo. Phylogenetic analysis of Erwinia species based on 16S rRNA gene sequences. Iternational Journal of Systematic Bacteriology. 1997; 47(4):1061-1067. https://doi.org/10.1099/00207713-47-4-1061
- Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. MEGA 5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Molecular Biology and Evolution. 2011; 28(10):2731-2739. https://doi.org/10.1093/molbev/msr121