UNIVERSAL AMPLIFICATION AND ANALYSIS OF PATHOGEN 16S RDNA FOR CLASSIFICATION AND IDENTIFICATION OF MYCOPLASMALIKE ORGANISMS

Regions representing about 80% of the 16S rDNA sequences of 40 mycoplasmalike organism (MLO) strains from North America, Asia, and Europe were amplified by polymerase chain reaction (PCR) using a primer pair designed on the basis of an MLO 16S rRNA gene. This primer pair detected every MLO examined from infected periwinkle (Catharanthus roseus) and some other plants. No PCR products were obtained in samples containing DNA extracted from healthy plants. The partial 16S rDNA sequences amplified from these various MLOs were compared through restriction fragment length polymorphism (RFLP) analyses. Based on similarity coefficients derived from RFLP analyses, these 40 MLOs could be classified into nine distinct 16S ribosomal RNA (16Sr) groups and 14 subgroups, including five groups that coincide with MLO strain clusters previously delineated on the basis of dot hybridization analysis using randomly cloned chromosomal DNA probes. Type MLO strains designated for each group and subgroup were as follows: 16SrI-A, tomato big bud; 16SrI-B, Maryland aster yellows; 16SrI-C, clover phyllody; 16SrI-D, paulownia witches'-broom; 16SrI-E, blueberry stunt; 16SrII, peanut witches'-broom; 16SrIII-A, Canada peach X; 16SrIII-B, clover yellow edge; 16SrIV, palm lethal yellowing; 16SrV, elm yellows; 16SrVI, clover proliferation; 16SrVII, ash yellows; 16SrVIII, loofah witches'-broom; and 16SrIX, pigeon pea witches-broom.