Aster yellows phytoplasmas are phytopathogenic mollicutes transmitted mainly by the leafhopper Macrosteles quadrilineatus Forbes. The 800 kb genome of the aster yellows phytoplasma subgroup A strain, aster yellows witches broom (AY-WB), has been sequenced near completion. The goals of this study are to characterize insertion sequences (ISs) in the AY-WB genome and to determine whether ISs can be used as markers to distinguish aster yellow phytoplasma isolates. Eleven AY-WB ISs were selected for further characterization. The organization and sequence of the 11 AY-WB ISs appear to be most similar to IS3 family insertion elements that have been characterized from the animal and human pathogenic mollicutes, the mycoplasmas as well. Seven ISs sequences of AY-WB ISs contain two adjacent open reading frames (ORFs), ORFA and ORFB, of which all but one are flanked by imperfect repeats (IRs) and/or direct repeats (DRs). In all seven ISs a frameshifting event may occur that results in ORFA and B fusion protein, which is the active transposase. However, only ISs ISAYWB-2 and ISAYWB-7 have a –1 frameshifting structure that is believed to result relatively frequently in synthesis of the ORFAB fusion protein. One AY-WB IS contains three ORFs due to a 39 bp gap in ORFB, and three AY-WB ISs contain single ORFs and lack IRs and DRs. Primers were designed on sequences flanking the ORFs and IRs, and/or between the IRs. Subsequent PCR of DNA of subgroup A and B aster yellows phytoplasma strains showed that ISs can be used for aster yellows phytoplasma isolate identification.
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