Open AccessLigation‐mediated PCR, a fast and reliable technique for insertion sequence‐based typing of Xanthomonas citri pv. citri
Author(s) -
Bui Thi Ngoc Lan,
Vernière Christian,
Belasque José,
Vital Karine,
Boutry Sébastien,
Gagnevin Lionel,
Pruvost Olivier
Publication year - 2008
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2008.01331.x
Subject(s) - xanthomonas citri , biology , restriction enzyme , insertion sequence , polymerase chain reaction , citrus canker , xanthomonas , genetics , microbiology and biotechnology , pathogen , dna , genome , gene , bacteria , transposable element
Asiatic citrus canker, caused by Xanthomonas citri pv. citri , is a major disease threatening citrus crops throughout the world. The most common methods for strain differentiation of this pathogen are repetitive element sequence‐based PCR (rep‐PCR) and pulsed field gel electrophoresis (PFGE), using rare‐cutting restriction enzyme analysis. We developed a ligation‐mediated PCR targeting three insertion sequences (IS‐LM‐PCR) present as several copies in the genome of the fully sequenced strain 306 of X. citri pv. citri . This technique amplifies DNA fragments between an insertion sequence element and an MspI restriction site. The analysis of strains can be conducted within 24 h, starting from very small amounts of bacterial DNA, which makes IS‐LM‐PCR much less labor‐intensive than PFGE. We used IS‐LM‐PCR to analyze a collection of 66 strains of X. citri pv. citri from around the world. The overall reproducibility of IS‐LM‐PCR reached 98% in this data set and its discriminatory power was markedly superior than rep‐PCR. We suggest that IS‐LM‐PCR could be used for the global surveillance of nonepidemiologically related strains of X. citri pv. citri .