|HUANG, H - University Of South Florida|
Submitted to: CDFA Pierce's Disease Control Program Research Symposium
Publication Type: Abstract Only
Publication Acceptance Date: 12/13/2011
Publication Date: 12/13/2011
Citation: Chen, J., Huang, H. 2011. Small RNAs in Xylella fastidiosa and their epidemiological applications. CDFA Pierce's Disease Control Program Research Symposium. p. 90.
Technical Abstract: Bacterial non-coding small RNAs (sRNAs) have attracted considerable attention due to their roles in regulating numerous cellular processes including survival, adaptation and pathogenesis. Sequence variation in sRNA genes reflect a previously unrecognized source of genomic diversity in bacteria. Xylella fastidiosa is an important bacterial pathogen causing many economically important diseases such as almond leaf scorch, citrus variegated chlorosis and Pierce’s disease of grapevine. Little is known about sRNAs in this bacterium. Therefore, a research project was initiated to search for sRNAs in X. fastidiosa. The complete genome sequences of four X. fastidiosa strains (9a5c, M12, M23, and Temecula1) respresenting three X. fastidiosa subspecies were selected and scanned for sRNA genes with established computer programs. Candidate sRNA genes were identified in all of the four X. fastidiosa strains (46 in strain 9a5c, 50 in strain M12, 49 in strain M23, and 47 in strain Temecula1). Candidate sRNA genes ranged in size from 40 to 350 bp. Expression of sRNA genes was proved using a procedure involving quantitative reverse transcriptase PCR (qRT-PCR) by comparing different X. fastidiosa strains cultured under the same condition or the same bacterial strain cultured under different conditions. BLAST analysis showed that 34 sRNA genes were shared by all three X. fastidiosa subspecies. To test for epidemiological application of variable sRNA genes, four sRNA genes in strain M23 were selected to design PCR primers. A total of 22 different bacterial strains were cultured in PW broth at 28 ºC for 14 days. DNA was extracted and used as templates for RT-PCR with the four sRNA primer sets. Both inter- and intra- subspecies variation of X. fastidiosa strains was observed.