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Title: CLASSIFICATION OF PHYTOPLASMAS BASED ON COMPUTER-SIMULATED RESTRICTION FRAGMENT LENGTH POLYMORPHISM ANALYSIS OF 16S RRNA GENE SEQUENCES

Author
item WEI, WEI - PEOPLES REPUBLIC OF CHINA
item Zhao, Yan
item Lee, Ing Ming
item Davis, Robert
item SUN, QINGRONG - PEOPLES REPUBLIC OF CHINA

Submitted to: International Organization for Mycoplasmology
Publication Type: Abstract Only
Publication Acceptance Date: 4/5/2006
Publication Date: 4/5/2006
Citation: Wei, W., Zhao, Y., Lee, I., Davis, R.E., Sun, Q. 2006. Classification of phytoplasmas based on computer-simulated restriction fragment length polymorphism analysis of 16S rRNA gene sequences. International Organization for Mycoplasmology. P. 105.

Interpretive Summary:

Technical Abstract: Phytoplasmas are cell wall-less pathogenic bacteria that cause numerous plant diseases. Due to the inability to culture phytoplasmas in vitro, it is impossible to differentiate and classify phytoplasmas by the traditional methods that are applied to cultured prokaryotes. To date, restriction fragment length polymorphism (RFLP) analysis of highly conserved 16S rRNA gene sequence has served as a primary method for differentiation and classification of phytoplasmas into groups and subgroups according to similarity coefficients [Lee et al., IJSB (1998) 48, 1153-1169]. However, RFLP analysis requires expensive multiple restriction enzyme digestions and subsequent polyacrylamide gel electrophoresis of DNA fragments. Furthermore, the number of groups and subgroups continues to expand, and comparative visual analysis of RFLP patterns is consequently becoming more difficult. To facilitate the implementation and expansion of the existing 16S rRNA gene sequence-based phytoplasma classification scheme, we developed a computer-simulated RFLP analysis method, which allows rapid determination of the classification status of any phytoplasma under investigation. Over 700 publicly available phytoplasma 16S rRNA gene sequences were used in this study. The sequences were aligned using the ClustalW algorithm, and the aligned 1.2 kb fragment was exported to software pDRAW32 for in silico restriction analysis and virtual gel plotting. Applying the same criteria (Lee et al., 1998), based on the simulated RFLP patterns and corresponding similarity coefficients, the phytoplasma strains were classified into 22 major groups and 77 subgroups. This study also determined the classification status of more than 250 previously unclassified phytoplasmas and established several new phytoplasma subgroups.