Lemon Lisbon (Citrus x limon cv. Lisbon) is an economically important citrus species grown in southern Iran. In 2018, several lemon Lisbon trees with typical symptoms of phytoplasms diseases were observed in the citrus orchards of Hormozgan province, Iran. The symptomatic plants showed witches’ broom, yellowing and narrowing leaves. To identify association of phytoplasma with the symptomatic plants, the total genomic DNA was extracted from 0.2 g midribs of four symptomatic and one asymptomatic plants using modified cetyltrimethylammonium bromide (CTAB) method. The samples were checked for phytoplasma DNA by direct-PCR using the phytoplasma universal primer pair P1/P7 and nested-PCR using primers P1/P7 (first round) followed by R16F2n/R16R2 (second round). Amplicons of the expected sizes (1800 and 1250 bp) were observed in direct and nested-PCR, respectively, in all symptomatic plants but not from symptomless plant or sterile distilled water as negative controls. A positive nested-PCR product was directly sequenced and edited using BioEdit (v. 5.0.9). BLAST search using sequenced contigs showed that the phytoplasma associated with lemon Lisbon had 100 % nucleotide identity with lime witches’ broom (LWB) phytoplasma (accession no: KU052827, MF071490 and LT558766), ‘Candidatus Phytoplasma aurantifolia’. Witches’ broom disease of lime is the most critical threats of limes and other susceptible citrus in southern Iran, Oman and the UAE, causing yield losses and destruction of the infected trees in a short time. To our knowledge, LWB associated phytoplasma is related to the Peanut witches' broom (16SrII) phytoplasma group of the Candidatus Phytoplasma. This report is the first study regarding of natural infection of lemon Lisbon trees by LWB phytoplasma in Iran and most probably in the world.