Multilocus Variable-Number-Tandem-Repeats Analysis (MLVA) distinguishes a clonal complex of Clavibacter michiganensis subsp. michiganensis strains isolated from recent outbreaks of bacterial wilt and canker in Belgium

BMC Microbiology

Paul De Vos²  Martine Maes³  Johan Van Vaerenbergh³  Pieter Stragier¹  Joanna Zaluga¹ 

[1] Laboratory of Microbiology, Department of Biochemistry and Microbiology, Ghent University, K.L. Ledeganckstraat 35, Gent, B-9000, Belgium;BCCM/LMG Bacteria collection - Laboratory of Microbiology Department of Biochemistry and Microbiology, Ghent University, K.L. Ledeganckstraat 35, Gent, B-9000, Belgium;Plant-Crop Protection, Institute for Agricultural and Fisheries Research, ILVO, Burg. Van Gansberghelaan 96, Merelbeke, B-9820, Belgium

关键词: Plant pathogen;    Bacterial wilt and canker;    Epidemiology;    Bacterial typing;    MLVA;    Clavibacter;   

Others  :  1143651
DOI  :  10.1186/1471-2180-13-126

 received in 2013-02-18, accepted in 2013-05-24,  发布年份 2013

【 摘 要 】

Background

Clavibacter michiganensis subsp. michiganensis (Cmm) causes bacterial wilt and canker in tomato. Cmm is present nearly in all European countries. During the last three years several local outbreaks were detected in Belgium. The lack of a convenient high-resolution strain-typing method has hampered the study of the routes of transmission of Cmm and epidemiology in tomato cultivation. In this study the genetic relatedness among a worldwide collection of Cmm strains and their relatives was approached by gyrB and dnaA gene sequencing. Further, we developed and applied a multilocus variable number of tandem repeats analysis (MLVA) scheme to discriminate among Cmm strains.

Results

A phylogenetic analysis of gyrB and dnaA gene sequences of 56 Cmm strains demonstrated that Belgian Cmm strains from recent outbreaks of 2010–2012 form a genetically uniform group within the Cmm clade, and Cmm is phylogenetically distinct from other Clavibacter subspecies and from non-pathogenic Clavibacter-like strains. MLVA conducted with eight minisatellite loci detected 25 haplotypes within Cmm. All strains from Belgian outbreaks, isolated between 2010 and 2012, together with two French strains from 2010 seem to form one monomorphic group. Regardless of the isolation year, location or tomato cultivar, Belgian strains from recent outbreaks belonged to the same haplotype. On the contrary, strains from diverse geographical locations or isolated over longer periods of time formed mostly singletons.

Conclusions

We hypothesise that the introduction might have originated from one lot of seeds or contaminated tomato seedlings that was the source of the outbreak in 2010 and that these Cmm strains persisted and induced infection in 2011 and 2012. Our results demonstrate that MLVA is a promising typing technique for a local surveillance and outbreaks investigation in epidemiological studies of Cmm.

【 授权许可】

   
2013 Zaluga et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 3.

【 参考文献 】

• [1]De León L, Siverio F, Lopez MM, Rodriguez A: Clavibacter michiganensis subsp. michiganensis, a seedborne tomato pathogen: healthy seeds are still the goal. Plant Dis 2011, 95(11):1328-1338.
• [2]EPPO: Clavibacter michiganensis subsp. michiganensis Diagnostics, PM7/42. Bulletin OEPP/EPPO 2005, 35:273-283.
• [3]Strider DL: Bacterial canker of tomato caused by Corynebacterium michiganense: a literature review and bibliography. Technical Bulletin North Carolina Agricultural Experiment Station; 1969:193.
• [4]Jahr H, Bahro R, Burger A, Ahlemeyer J, Eichenlaub R: Interactions between Clavibacter michiganensis and its host plants. Environ Microbiol 1999, 1(2):113-118.
• [5]Lamichhane JR, Balestra GM, Varvaro L: Severe Outbreak of Bacterial Canker Caused by Clavibacter michiganensis subsp. michiganensis on Tomato in Central Italy. Plant Dis 2011, 95(2):221-221.
• [6]De León L, Rodriguez A, Llop P, Lopez MM, Siverio F: Comparative study of genetic diversity of Clavibacter michiganensis subsp. michiganensis isolates from the Canary Islands by RAPD-PCR, BOX-PCR and AFLP. Plant Pathol 2009, 58(5):862-871.
• [7]Milijašević-Marčić S, Gartemann KH, Frohwitter J, Eichenlaub R, Todorović B, Rekanović E, Potočnik I: Characterization of Clavibacter michiganensis subsp. michiganensis strains from recent outbreaks of bacterial wilt and canker in Serbia. Eur J Plant Pathol 2012, 134:697-711.
• [8]Kawaguchi A, Tanina K, Inoue K: Molecular typing and spread of Clavibacter michiganensis subsp. michiganensis in greenhouses in Japan. Plant Pathol 2010, 59(1):76-83.
• [9]Bella P, Ialacci G, Licciardello G, La Rosa R, Catara V: Characterization of atypical Clavibacter michiganensis subsp. michiganensis populations in greenhouse tomatoes in Italy. J Plant Pathol 2012, 94(3):635-642.
• [10]Kleitman F, Barash I, Burger A, Iraki N, Falah Y, Sessa G, Weinthal D, Chalupowicz L, Gartemann K, Eichenlaub R: Characterization of a Clavibacter michiganensis subsp. michiganensis population in Israel. Eur J Plant Pathol 2008, 121(4):463-475.
• [11]Sahin F, Uslu H, Kotan R, Donmez M: Bacterial canker, caused by Clavibacter michiganensis subsp. michiganensis, on tomatoes in eastern Anatolia region of Turkey. Plant Pathol 2002, 51(3):399-399.
• [12]Cangelosi GA, Freeman RJ, Lewis KN, Livingston-Rosanoff D, Shah KS, Milan SJ, Goldberg SV: Evaluation of a high-throughput repetitive-sequence-based PCR system for DNA fingerprinting of Mycobacterium tuberculosis and Mycobacterium avium complex strains. J Clin Microbiol 2004, 42(6):2685-2693.
• [13]Bosch T, de Neeling A, Schouls L, Zwaluw K, Kluytmans J, Grundmann H, Huijsdens X: PFGE diversity within the methicillin-resistant Staphylococcus aureus clonal lineage ST398. BMC Microbiol 2010, 10(1):40. BioMed Central Full Text
• [14]Van Belkum A: Tracing isolates of bacterial species by multilocus variable number of tandem repeat analysis (MLVA). FEMS Immunol Med Microbiol 2006, 49(1):22-27.
• [15]Van Belkum A, Scherer S, Van Alphen L, Verbrugh H: Short-sequence DNA repeats in prokaryotic genomes. Microbiol Mol Biol Rev 1998, 62(2):275.
• [16]Harth-Chu E, Espejo RT, Christen R, Guzman CA, Hofle MG: Multiple-locus variable-number tandem-repeat analysis for clonal identification of Vibrio parahaemolyticus isolates by using capillary electrophoresis. Appl Environ Microbiol 2009, 75(12):4079-4088.
• [17]Lindstedt BA, Heir E, Gjernes E, Vardund T, Kapperud G: DNA fingerprinting of Shiga-toxin producing Escherichia coli O157 based on multiple-locus variable-number tandem-repeats analysis (MLVA). Ann Clin Microbiol Antimicrob 2003, 2(1):12. BioMed Central Full Text
• [18]Danin-Poleg Y, Cohen LA, Gancz H, Broza YY, Goldshmidt H, Malul E, Valinsky L, Lerner L, Broza M, Kashi Y: Vibrio cholerae strain typing and phylogeny study based on simple sequence repeats. J Clin Microbiol 2007, 45(3):736-746.
• [19]Heymans R, Schouls LM, van der Heide HG: Schim van der Loeff MF, Bruisten SM: Multiple-locus variable-number tandem repeat analysis of Neisseria gonorrhoeae. J Clin Microbiol 2011, 49(1):354-363.
• [20]van Cuyck H, Pichon B, Leroy P, Granger-Farbos A, Underwood A, Soullie B, Koeck J-L: Multiple-locus variable-number tandem-repeat analysis of Streptococcus pneumoniae and comparison with multiple loci sequence typing. BMC Microbiol 2012, 12(1):241. BioMed Central Full Text
• [21]Skuce RA, McCorry TP, McCarroll JF, Roring SMM, Scott AN, Brittain D, Hughes SL, Hewinson RG, Neill SD: Discrimination of Mycobacterium tuberculosis complex bacteria using novel VNTR-PCR targets. Microbiology 2002, 148(2):519-528.
• [22]Marsh JW, O’Leary MM, Shutt KA, Pasculle AW, Johnson S, Gerding DN, Muto CA, Harrison LH: Multilocus variable-number tandem-repeat analysis for investigation of Clostridium difficile transmission in hospitals. J Clin Microbiol 2006, 44(7):2558-2566.
• [23]Hidalgo Ă, Carvajal A, La T, Naharro G, Rubio P, Phillips ND, Hampson DJ: Multiple-locus variable-number tandem-repeat analysis of the swine dysentery pathogen, Brachyspira hyodysenteriae. J Clin Microbiol 2010, 48(8):2859-2865.
• [24]Le Fleche P, Hauck Y, Onteniente L, Prieur A, Denoeud F, Ramisse V, Sylvestre P, Benson G, Ramisse F, Vergnaud G: A tandem repeats database for bacterial genomes: application to the genotyping of Yersinia pestis and Bacillus anthracis. BMC Microbiol 2001, 1(1):2. BioMed Central Full Text
• [25]Li Y, Cui Y, Hauck Y, Platonov ME, Dai E, Song Y, Guo Z, Pourcel C, Dentovskaya SV, Anisimov AP: Genotyping and phylogenetic analysis of Yersinia pestis by MLVA: insights into the worldwide expansion of Central Asia plague foci. PLoS One 2009, 4(6):e6000.
• [26]Zhao WJ, Chen HY, Zhu SF, Xia MX, Tan TW: One-step detection of Clavibacter michiganensis subsp. michiganensis in symptomless tomato seeds using a Taqman probe. J Plant Pathol 2007, 89(3):349-351.
• [27]Gironde S, Manceau C: Housekeeping Gene Sequencing and Multilocus Variable-Number Tandem-Repeat Analysis To Identify Subpopulations within Pseudomonas syringae pv. maculicola and Pseudomonas syringae pv. tomato That Correlate with Host Specificity. Appl Environ Microbiol 2012, 78(9):3266-3279.
• [28]Coletta-Filho HD, Takita MA, De Souza AA, Aguilar-Vildoso CI, Machado MA: Differentiation of strains of Xylella fastidiosa by a variable number of tandem repeat analysis. Appl Environ Microbiol 2001, 67(9):4091-4095.
• [29]Wang DY, Hadj-Henni L, Thierry S, Arna P, Chermette R, Botterel F, Hadrich I, Makni F, Ayadi A, Ranque S: Simple and Highly Discriminatory VNTR-Based Multiplex PCR for Tracing Sources of Aspergillus flavus Isolates. PLoS One 2012, 7(9):e44204.
• [30]Bergsma-Vlami M, Martin W, Koenraadt H, Teunissen H, Pothier J, Duffy B, van Doorn J: Molecular typing of Dutch isolates of Xanthomonas arboricola pv. pruni isolated from ornamental cherry laurel. J Plant Pathol 2012, 94(1):S1. 29-S21. 35.
• [31]Bui Thi Ngoc L, Vernire C, Jarne P, Brisse S, Guerin F, Boutry S, Gagnevin L, Pruvost O: From local surveys to global surveillance: three high-throughput genotyping methods for epidemiological monitoring of Xanthomonas citri pv. citri pathotypes. Appl Environ Microbiol 2009, 75(4):1173-1184.
• [32]Zaluga J, Heylen K, Van Hoorde K, Hoste B, Van Vaerenbergh J, Maes M, De Vos P: GyrB sequence analysis and MALDI-TOF MS as identification tools for plant pathogenic Clavibacter. Syst Appl Microbiol 2011, 34(6):400-407.
• [33]Jacques MA, Durand K, Orgeur G, Balidas S, Fricot C, Bonneau S, Quillévéré A, Audusseau C, Olivier V, Grimault V: Phylogenetic analysis and polyphasic characterization of Clavibacter michiganensis strains isolated from tomato seeds reveal that non-pathogenic strains are distinct from C. michiganensis subsp. michiganensis. Appl Environ Microbiol 2012, 78(23):8388-8402.
• [34]ISF: Methods for the detection of Clavibacter michiganensis ssp michiganensis on tomato seeds Version 4. 2011. http://www.worldseed.org/isf/ishi_vegetable.html webcite
• [35]Jansing H, Rudolph K: Physiological capabilities of Clavibacter michiganensis subsp. sepedonicus and development of a semi-selective medium. Zeitschrift Fur Pflanzenkrankheiten Und Pflanzenschutz-Journal of Plant Diseases and Protection 1998, 105(6):590-601.
• [36]Pitcher D, Saunders N, Owen R: Rapid extraction of bacterial genomic DNA with guanidium thiocyanate. Lett Appl Microbiol 1989, 8(4):151-156.
• [37]Waleron M, Waleron K, Kamasa J, Przewodowski W, Lojkowska E: Polymorphism analysis of housekeeping genes for identification and differentiation of Clavibacter michiganensis subspecies. Eur J Plant Pathol 2011, 131(2):341-354.
• [38]Schneider KL, Marrero G, Alvarez AM, Presting GG: Classification of plant associated bacteria using RIF, a computationally derived DNA marker. PLoS One 2011, 6(4):e18496.
• [39]Thompson J, Higgins D, Gibson T: CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 1994, 22(22):4673.
• [40]Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ: Basic local alignment search tool. J Mol Biol 1990, 215(3):403-410.
• [41]Tamura K, Dudley J, Nei M, Kumar S: MEGA4: Molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol Biol Evol 2007, 24(8):1596-1599.
• [42]Tamura K, Nei M: Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol Biol Evol 1993, 10(3):512-526.
• [43]Librado P, Rozas J: DnaSP v5: a software for comprehensive analysis of DNA polymorphism data. Bioinformatics 2009, 25(11):1451-1452.
• [44]Hunter PR, Gaston MA: Numerical index of the discriminatory ability of typing systems: an application of Simpson’s index of diversity. J Clin Microbiol 1988, 26(11):2465-2466.
• [45]Gelfand Y, Rodriguez A, Benson G: TRDB - the tandem repeats database. Nucleic Acids Res 2007, 35(suppl 1):D80-D87.
• [46]Rozen S, Skaletsky H: Primer3 on the WWW for general users and for biologist programmers. Methods Mol Biol 2000, 132(3):365-386.
• [47]Simpson EH: Measurement of diversity. Nature: Nature; 1949.
• [48]Nazari F, Niknam GR, Ghasemi A, Taghavi SM, Momeni H, Torabi S: An investigation on strains of Clavibacter michiganensis subsp. michiganensis in north and north west of Iran. J Phytopathol 2007, 155(9):563-569.
• [49]Klevytska AM, Price LB, Schupp JM, Worsham PL, Wong J, Keim P: Identification and characterization of variable-number tandem repeats in the Yersinia pestis genome. J Clin Microbiol 2001, 39(9):3179-3185.
• [50]Sobral D, Schwarz S, Bergonier D, Brisabois A, Feßler AT, Gilbert FB, Kadlec K, Lebeau B, Loisy-Hamon F, Treilles M: High Throughput Multiple Locus Variable Number of Tandem Repeat Analysis (MLVA) of Staphylococcus aureus from Human. Animal and Food Sources. PLoS One 2012, 7(5):e33967.
• [51]Call DR, Orfe L, Davis MA, Lafrentz S, Kang M-S: Impact of compounding error on strategies for subtyping pathogenic bacteria. Foodborne Pathog Dis 2008, 5(4):505-516.
• [52]Gulati P, Varshney R, Virdi J: Multilocus variable number tandem repeat analysis as a tool to discern genetic relationships among strains of Yersinia enterocolitica biovar 1A. J Appl Microbiol 2009, 107(3):875-884.
• [53]Broschat S, Call D, Davis M, Meng D, Lockwood S, Ahmed R, Besser T: Improved identification of epidemiologically related strains of Salmonella enterica by use of a fusion algorithm based on pulsed-field gel electrophoresis and multiple-locus variable-number tandem-repeat analysis. J Clin Microbiol 2010, 48(11):4072-4082.
• [54]Domenech P, Barry C 3rd, Cole ST: Mycobacterium tuberculosis in the post-genomic age. Curr Opin Microbiol 2001, 4(1):28.
• [55]Pourcel C, Minandri F, Hauck Y, D’Arezzo S, Imperi F, Vergnaud G, Visca P: Identification of variable-number tandem-repeat (VNTR) sequences in Acinetobacter baumannii and interlaboratory validation of an optimized multiple-locus VNTR analysis typing scheme. J Clin Microbiol 2011, 49(2):539-548.
• [56]Kremer K, Arnold C, Cataldi A, Gutierrez MC, Haas WH, Panaiotov S, Skuce RA, Supply P, van der Zanden AGM, van Soolingen D: Discriminatory power and reproducibility of novel DNA typing methods for Mycobacterium tuberculosis complex strains. J Clin Microbiol 2005, 43(11):5628-5638.