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Faghihi, M. M., Bagheri, A. N., Bahrami, H. R., Hasanzadeh, H., Rezazadeh, R., Siampour, M., Samavi, S., Salehi, M., and Izadpanah, K. 2011. Witches'-broom disease of lime affects seed germination and seedling growth but is not seed transmissible. Plant Dis. 95:419-422.Witches'-broom disease of lime (WBDL), caused by 'Candidatus Phytoplasma aurantifolia', has devastated many Mexican lime orchards and is currently a threat to lime production in neighboring provinces of southern Iran. Several reports have suggested transmission and spread of WBDL phytoplasma through the seed of infected plants. In the present study, claims of seed transmission of this phytoplasma were examined. Fruit were collected from infected trees in the infested areas of Minab (Hormozgan Province) and from symptomless trees in noninfested areas. Lime seed from symptomless and witches'-broomaffected trees were sown in separate beds in an insect-proof screenhouse and the resulting seedlings were examined for phytoplasmal infection. Leaf, stem, and root samples were collected from both groups of seedlings every 3 months for 2 years and tested for WBDL phytoplasma using direct and nested polymerase chain reaction (PCR).Repeated PCR tests on the seedlings did not reveal the presence of phytoplasmal DNA. Likewise, symptoms of the disease were not observed on these seedlings after 2 years. PCR assays detected the phytoplasma in coats of some seed from infected trees; however, no excised embryos were positive for the phytoplasma. All positive PCR results were confirmed by restriction fragment length polymorphism assay.One-year-old seedlings derived from seed of noninfected plants appeared more vigorous in terms of height, number of leaves, and fresh weight of shoot compared with those from infected trees. The germination percentage, mean daily germination, peak value, and germination value were significantly higher for seed of fruit from noninfected trees and seed from fruit on asymptomatic branches of infected trees than those from fruit on symptomatic branches of infected trees.Witches'-broom disease of lime (WBDL), caused by 'Candidatus Phytoplasma aurantifolia' was found first in a small area in southern Iran in 1997 (17). Since then, WBDL has spread to many areas, devastating Mexican lime plantations in major lime-growing provinces of Hormozgan, Sistan-Baluchistan, and Kerman (16). In the field, the phytoplasma is naturally spread by the vector leafhopper Hishimonus phycitis (18). The pathogen has been experimentally transmitted to several species of citrus by graft inoculation, and to periwinkle and a number of solanaceous plants such as tomato, eggplant, and tobacco by dodder or graft inoculation (19)(20)(21). Several studies have focused on the detection of WBDL phytoplasma in various parts of the lime seed. were the first to suggest the possibility of seed transmission of WBDL phytoplasma. Later, Khan (9) used the polymerase chain reaction (PCR) assay to study seed transmission of WBDL phytoplasma in 6-month-old seedlings. Based on this repo...

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Somatic hybridization of mango via protoplast fusion was attempted at cultivar level. Enzymatically isolated protoplasts from leaves of greenhouse-grown seedlings of cvs. 'Tommy Atkins', 'Keitt' and 'Haden' and from proembryonic masses (PEMs) of cv. 'Kensington Pride' were used. Protoplasts were fused by polyethylene glycol (PEG), embedded in Ca-alginate beads and cultured in shallow liquid culture on shaker (30 rpm). After 4 weeks, Ca-alginate beads were depolymerized and released microcolonies of PEMs were plated onto the solid culture media. After two consecutive subcultures, fast growing large clumps of PEMs were picked up and cultured as PEMs line for analyses. Flow cytometry analysis of 242 PEMs lines revealed 41 tetraploid lines. DNA fi ngerprinting of the regenerated embryos from the tetraploid lines showed that only four lines were somatic hybrids, all resulting from 'Haden' + 'Kensington Pride' protoplast fusions. By contrast, the tetraploid lines from 'Keitt' + 'Kensington Pride' and 'Tommy Atkins' + 'Kensington Pride' were autotetraploids. Root-tip chromosome counts on resulting germinated cotyledonary embryos confi rmed that somatic hybrid embryo lines had a chromosome number of 2n=4x=80 compared to diploid parents (2n=2x=40). Of 50 defl asked somatic-hybrid, in vitro plantlets with true leaves only 3 plantlets formed the healthy apical bud (meristem) in the soil and grew normally.