Phytoplasma-related symptoms such as big bud, witches' broom, phyllody, virescence, flower proliferation, little leaves, yellowing, and stem fasciation were observed in 9 plant species during a field survey in Fayoum Governorate, Egypt, in 2021 and 2022. Disease incidence on various crops ranged from 1 to 21.3% in 2021, with the percentage rising in 2022. In 2022, the highest incidence percentage (23 %( was recorded on sesame while on cowpea the lowest percentage of incidence was recorded being (1.2%). A nested PCR assay employing the universal primer pair P1/P7 followed by R16F2n/R16R2 was used to detect the presence of phytoplasmas by amplifying a segment of the phytoplasma 16S rRNA gene. The resulting amplicons were purified and sequenced. The sequences were deposited in GenBank. Sequence comparison, phylogenetic study, and virtual RFLP analysis of 16S rRNA indicated the presence of diverse phytoplasma ribosomal groups and subgroups infecting different crops (16SrVI-A, 16SrIII-A, 16SrII-D, 16SrII-A, 16SrI-A, and 16SrI-B). The 16SrVI-A phytoplasmas were detected in symptomatic tomato plants, the 16SrII-D phytoplasmas subgroup was identified in sesame, faba bean, and cowpea exhibiting phyllody symptoms, the 16SrII-A subgroup was detected in cauliflower. The 16SrIII-A and 16SrI-A phytoplasmas subgroups were identified in alfalfa witches' broom and chamomile, respectively for the first time in Egypt. Moreover, subgroup 16SrI-B was detected for the first time in Egypt in chrysanthemum and cabbage. This study showed the genetic diversity of phytoplasma strains infecting a variety of crops in Fayoum Governorate.