Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 5/12/2006
Publication Date: 7/8/2006
Citation: Mello, A.F.S., Yokomi, R.K., Melcher, U., Chen, J.C., Fletcher. J. 2006. Molecular techniques to assess genetic diversity among isolates of Spiroplasma citri [abstract]. 16th International Congress International Organization for Mycoplasmology (IOM), St John's College Cambridge, UK, July 9-14, 2006.
Technical Abstract: Spiroplasma citri, a phloem restricted pathogen transmitted by leafhoppers, causes stubborn disease in citrus. Production losses attributed to this disease in California have become a growing concern over the past decade. One possible explanation for the re-emergence of stubborn disease is the introduction or emergence of one or more new pathogen strains. The aim of this study was to evaluate repetitive extragenic palindromic elements (rep) and random amplified polymorphic DNA (RAPD) as discriminative techniques to asses the genetic diversity among isolates and thereby evaluate the pathogen’s evolutionary history and specificity to hosts and locations. Ten S. citri isolates from Iran, Israel, Morocco and different regions of the U.S. were cultured in LD8 medium. Culture suspensions, or extracted and purified DNA, were used as PCR templates. Three rep primers (BOX, ERIC and REP) and five RAPD primers were used in PCR with three replications. Use of purified DNA templates provided more discriminatory band patterns than did the bacterial cultures. BOX was the only rep-PCR primer to produce differential band patterns in the evaluated isolates. One RAPD primer revealed a strain-differential 520 bp band in both cultures and purified DNA, and a second primer yielded two bands that discriminated among the isolates only when purified DNA was used. One of these bands (2100 bp) occurred in 5 isolates and the other (1000 bp) was unique to a single isolate. These data suggest that there is genetic variability among different populations of S. citri and that some RAPD primers and the rep-BOX are useful to evaluate such diversity.