1. Comparative genomics to develop a specific multiplex PCR assay for detection of Clavibacter michiganensis
- Author
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Shree Prasad Thapa, Michael O’Leary, Gitta Coaker, Marie-Agnès Jacques, Robert L. Gilbertson, Department of Plant Pathology, University of Kentucky, Institut de Recherche en Horticulture et Semences (IRHS), Université d'Angers (UA)-AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and GC, SP, and RG were supported by grants from the California Tomato 22 Research Institute awarded to GC and RG (091). GC and SP were supported by a grant 23 from USDA-NIFA (2015-67013-23082, awarded to GC) and grants awarded by the Page 1 of 56 2 24 California Tomato Research Institute and the California Department of Food and 25 Agriculture (17-0275-047-SC).
- Subjects
[SDV.SA]Life Sciences [q-bio]/Agricultural sciences ,0106 biological sciences ,[SDV.OT]Life Sciences [q-bio]/Other [q-bio.OT] ,génome végétal ,Plant Science ,lutte contre les bactéries ,01 natural sciences ,Tomato ,Multiplex PCR ,Diagnostic ,Clavibacter michiganensis ,Comparative 19 genomics ,Microbiology ,03 medical and health sciences ,Plasmid ,résistance aux agents pathogènes ,[SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN] ,Multiplex polymerase chain reaction ,medicine ,protection des plantes ,Pathogen ,cartographie chromosomique ,chancre bactérien ,030304 developmental biology ,2. Zero hunger ,Comparative genomics ,Canker ,0303 health sciences ,Vegetal Biology ,biology ,Bacterial wilt ,Microbiology and Parasitology ,food and beverages ,16S ribosomal RNA ,biology.organism_classification ,medicine.disease ,Microbiologie et Parasitologie ,Agricultural sciences ,[SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology ,pathogène des semences ,prolifération ,Agronomy and Crop Science ,Biologie végétale ,Sciences agricoles ,010606 plant biology & botany ,Autre (Sciences du Vivant) - Abstract
Clavibacter michiganensis is a Gram-positive bacterial pathogen that proliferates in the xylem vessels of tomato, causing bacterial wilt and canker symptoms. Accurate detection is a crucial step in confirming outbreaks of bacterial canker and developing management strategies. A major problem with existing detection methods are false-positive and -negative results. Here, we report the use of comparative genomics of 37 diverse Clavibacter strains, including 21 strains sequenced in this study, to identify specific sequences that are C. michiganensis detection targets. Genome-wide phylogenic analyses revealed additional diversity within the genus Clavibacter. Pathogenic C. michiganensis strains varied in plasmid composition, highlighting the need for detection methods based on chromosomal targets. We utilized sequences of C. michiganensis-specific loci to develop a multiplex PCR-based diagnostic platform using two C. michiganensis chromosomal genes (rhuM and tomA) and an internal control amplifying both bacterial and plant DNA (16s ribosomal RNA). The multiplex PCR assay specifically detected C. michiganensis strains from a panel of 110 additional bacteria, including other Clavibacter spp. and bacterial pathogens of tomato. The assay was adapted to detect the presence of C. michiganensis in seed and tomato plant materials with high sensitivity and specificity. In conclusion, the described method represents a robust, specific tool for detection of C. michiganensis in tomato seed and infected plants.
- Published
- 2019
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