Characterization of insertions of IS476 and two newly identified insertion sequences, IS1478 and IS1479, in
J H Chen1, Y Y Hsieh, S L Hsiau
1Institute of Molecular Biology, National Chung Hsing University, Taichung 40227, Taiwan, Republic of China. jhchen@dragon.nchu.edu.tw
Abstract:
Thirty-two plasmid insertion mutants were independently isolated from two strains of Xanthomonas campestris pv. campestris in Taiwan. Of the 32 mutants, 14 (44%), 8 (25%), and 4 (12%) mutants resulted from separate insertions of an IS3 family member, IS476, and two new insertion sequences (IS), IS1478 and IS1479. While IS1478 does not have significant sequence homology with any IS elements in the EMBL/GenBank/DDBJ database, IS1479 demonstrated 73% sequence homology with IS1051 in X. campestris pv. dieffenbachiae, 62% homology with IS52 in Pseudomonas syringae pv. glycinea, and 60% homology with IS5 in Escherichia coli. Based on the predicted transposase sequences as well as the terminal nucleotide sequences, IS1478 by itself constitutes a new subfamily of the widespread IS5 family, whereas IS1479, along with IS1051, IS52, and IS5, belongs to the IS5 subfamily of the IS5 family. All but one of the IS476 insertions had duplications of 4 bp at the target sites without sequence preference and were randomly distributed. An IS476 insertion carried a duplication of 952 bp at the target site. A model for generating these long direct repeats is proposed. Insertions of IS1478 and IS1479, on the other hand, were not random, and IS1478 and IS1479 each showed conservation of PyPuNTTA and PyTAPu sequences (Py is a pyrimidine, Pu is a purine, and N is any nucleotide) for duplications at the target sites. The results of Southern blot hybridization analysis indicated that multiple copies of IS476, IS1478, and IS1479 are present in the genomes of all seven X. campestris pv. campestris strains tested and several X. campestris pathovars.
Insights
Researchers identified new insertion sequences (IS) IS1478 and IS1479 in Xanthomonas campestris pv. campestris. These new elements, along with IS476, are widespread in Xanthomonas campestris strains and exhibit distinct insertion mechanisms.
Area of Science:
- Microbiology
- Genetics
- Molecular Biology
Background:
- Xanthomonas campestris pv. campestris causes black rot in crucifers.
- Insertion sequences (IS) are mobile genetic elements that can influence bacterial genome evolution and pathogenicity.
- Understanding IS elements in plant pathogens is crucial for disease management.
Purpose of the Study:
- To characterize novel insertion sequences (IS) identified in Xanthomonas campestris pv. campestris.
- To investigate the insertion mechanisms and distribution of these IS elements within Xanthomonas species.
Main Methods:
- Isolation and characterization of plasmid insertion mutants.
- Sequence analysis of novel IS elements (IS1478, IS1479) and known elements (IS476).
- Southern blot hybridization to assess the genomic distribution of IS elements.
Main Results:
- Four types of IS elements were identified: IS3 family member, IS476, IS1478, and IS1479.
- IS1478 represents a new subfamily of the IS5 family; IS1479 belongs to the IS5 subfamily.
- IS476 insertions were random with 4 bp duplications, while IS1478 and IS1479 showed non-random insertions with specific target site sequences.
- IS476, IS1478, and IS1479 are present in multiple copies across various Xanthomonas campestris strains and pathovars.
Conclusions:
- Discovery of two novel insertion sequences, IS1478 and IS1479, in Xanthomonas campestris pv. campestris.
- Elucidation of distinct insertion preferences and mechanisms for different IS elements.
- Demonstration of the widespread presence and potential mobility of these IS elements in Xanthomonas species, highlighting their role in bacterial evolution.
