Witches-broom disease of lime (WBDL) caused by 'Candidatus Phytoplasma aurantifolia' is a devastating disease in the Sultanate of Oman, United Arab Emirates, and southern Iran. The disease has been experimentally transmitted to lime trees by the leafhopper Hishimonus phycitis and to lime seedlings by grafting. However, the natural transmission of phytoplasma to lime seedlings by the vector has not been reported. Based on the previous studies, although the disease has been transmitted to Bakraee seedlings under greenhouse condition, all efforts to transmit the disease by a vector to lime seedlings grown in greenhouse have been failed. Considering that the transmission of witches' broom phytoplasma to 15-20 year old trees grown under semi-natural environment was successful and given to the fact that lime is the main preferred host of this phytoplasma, the current study was carried out to re-examine the transmission of witches' broom phytoplasma to lime seedlings by H. phycitis. The leafhopper nymphs and adults were collected in a WBDL-infected lime orchard by D-Vac aspirator from Roudan (Hormozgan Province) in May 2017 and some of them were tested for phytoplasma infection. Fifteen healthy one-year old seedlings were provided from Minab, Hormozgan Province. Total DNA were extracted from plants and leafhoppers according to Doyle (1990) and Reineke et al., 1998), respectively. Of more than 100 leafhopper samples tested, at least 90% were positive for the phytoplasma by PCR using specific primer. Additional field-collected leafhoppers were caged (five cages each containing five leafhoppers) on Mexican lime seedlings in the greenhouse in Bandar Abbas (Hormozgan Province). In addition, almost 100 H. phycitis (some examined by PCR) collected from non-infected trees in disease-free areas were similarly on five healthy Mexican lime seedlings as negative control. Three months after inoculation, 5 of 10 inoculated seedlings showed typical symptoms of WBDL, including bud proliferation, general chlorosis, and stunting. Symptomatic plants were strongly positive in PCR assays using specific primer but not form negative control. Amplified R16f2n/R16r2 primers PCR products (1,250 bp) from experimentally vector challenged Mexican lime seedlings were sequenced bidirectional. BLAST analysis of the 16S rDNA sequences revealed that the phytoplasma associated with lime seedlings shared 100% identity with 'Candidatus Phytoplasma aurantifolia'. To our knowledge, this is the first report of natural transmission of Ca. P. aurantifolia to Mexican lime seedlings by H. phycitis which can be a serious alarm to the lime seedlings producers and new re-cultivated lime orchards of southern Iran. Since there is many lime genotypes with different susceptibility to the phytoplasma disease in southern Iran and re-planting of Mexican lime in southern Iran has been started currently, use of resistant or tolerant lime genotypes for re-cultivation programs is strongly suggested, and otherwise, a new phytoplasma epidemic outbreak is not unexpected.