USE OF MYCOPLASMALIKE ORGANISM (MLO) GROUP-SPECIFIC OLIGONUCLEOTIDE PRIMERS FOR NESTED-PCR ASSAYS TO DETECT MIXED-MLO INFECTIONS IN A SINGLE HOST-PLANT

Oligonucleotide primer pairs R16(I)FI/R1, R16(III)F2/R1, and R16(V)F1/ RI for polymerase chain reactions (PCRs) were designed on the basis of mycoplasmalike organism (MLO) 16S rRNA sequences. The primer pair R16 (I)F1/R1 specifically initiated amplification of 16S rDNA sequences am ong MLO strains in the MLO 16S rRNA group I, which includes aster yell ows MLO and related strains; R16(III)F2/R1 specifically initiated ampl ification in the MLO 16S rRNA group III, which includes peach X-diseas e MLO and related strains; and R16(V)F1/R1 specifically initiated ampl ification in the MLO 16S rRNA group V, which includes elm yellows MLO and related strains. None of the primer pairs initiated amplification of 16S rDNA sequences from MLO strains in other groups or from other p rokaryotes, including animal Mollicutes and plant pathogenic bacteria. An MLO group-specific primer pair allows sensitive detection and simu ltaneous classification of specific MLO strains from plant and insect sources. Nested-PCR assays using the universal primer pair R16F2/R2 an d a group-specific primer pair further increased sensitivity in MLO de tection. These specially designed assay procedures allowed for the fir st time detection of a secondary, cryptic MLO(s) associated with a sin gle host plant.