Detection and molecular identification of phytoplasma associated with Gazania in Egypt
Detection and molecular identification of phytoplasma associated with Gazania in Egypt
Shimaa M. Gad1, Ahmed A. Kheder1, Mohamed A. Awad 2
ABSTRACT
Background: Phytoplasmas a wall-less microorganism belongs to Class Mollicutes cause diseases in
several commercial ornamental plants, leads to serious economic losses all over the world. During
2017-2018, gazania showing phyllody, yellowing, proliferation, virescence and little leaf symptoms
was observed Giza, Egypt.
Objective: The current work aims the detection and molecular identification of phytoplasma infecting
Gazania in Egypt.
Methods: Phytoplasma disease was detected and isolated from naturally infected gazania plants during
surveys in flower nurseries and open filed in Giza governorate, it was transmitted from naturally
infected Gazania to healthy periwinkle and other ornamental plants by dodder (Cuscuta reflexa ), and
the leaf hopper (Empoasca decipiens). Free hand sections stained with Diene's stain was used to detect
and differentiate the phloem tissues of leaf sections from infected gazania and periwinkle plants.
Transmission Electron Microscopy (TEM) revealed the presence of phytoplasma in the sieve tubes and
parenchyma cells of leaf midribs in infected plants. DNA extracted from symptomatic samples was
used as a template in nested polymerase chain reaction (nested-PCR) using universal primer pairs
P1/P7 and R16F2n/R16R2. Sequencing and phylogenetic analysis were performed to identify the
detected phytoplasmas.
Results: Phytoplasma was transmitted successfully from naturally infected gazania to healthy
ornamental plants by dodder and insect. Transmission electron microscopy (TEM) revealed that,
phytoplasma-like bodies were detected inside phloem, sieve tubes and parenchyma cells of leaf
midribs tissues in infected plants and ranging from 200 to 400nm in diameters. The 16SrRNA gene
from phytoplasma was amplified by nested-PCR assay and direct sequenced using specific primer
pairs. Phylogenetic tree was based on obtained sequences data.
Conclusion: The phytoplasma associated with gazania exhibiting phyllody, yellowing, proliferation,
virescence and little leaf symptoms was confirmed by the results of LM and TEM observations and
nested-PCR testing. Based on direct sequence date, phylogeny analysis, the associated phytoplasma
was classified as related to 16SrII group.
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July 2019
Vol. 6, Pages 1-101
