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    First report of 'candidatus phytoplasma trifolii' associated with leaf reddening and upright growth in pears (Pyrus communis l.)
    (Czech Academy of Agricultural Sciences, 2021) Usta, Mustafa; Güller, Abdullah; Sipahioğlu, Hikmet Murat
    The natural occurrence of 'Candidatus Phytoplasma trifolii' in pear trees (Pyrus communis Linnaeus) is reported here for the first time. In 2017, a total of thirty-five pear trees, two of them exhibiting leaf rolling along the mid-vein, reddening, bushy appearance, and upright growth symptoms were sampled in different locations in Van province, Turkey. The total deoxyribonucleic acid was extracted from symptomatic and asymptomatic plants. The purified DNA served as a template in nested polymerase chain reaction (nested-PCR) assays, performed to amplify 16S rRNA sequences using universal primer pairs (R16mF2/R16mR1 and R16F2n/R16R2). The resulting PCR products were then cloned into a pGEM T-Easy vector and sequenced bidirectionally. The phytoplasma strain, group, and subgroup identity were determined using the in silico restriction fragment length polymorphism (RFLP) analysis of the 16S ribosomal RNA-encoding gene sequences profiling with seventeen distinct restriction enzymes. Of the thirty-five pear samples, only two yielded 1 256 bp and 1 258 bp DNA fragments and were designated as Van-Pr3 (Acc. No. MH709141) and Van-Pr4 (Acc. No. MH730561), respectively. Based on the in silico virtual RFLP pattern analysis of the 16S rRNA se-quences, we confirmed the presence of 'Ca. P. trifolii' belonging to the clover proliferation group and both identified phytoplasmas were identical with the similarity coefficient of 1.00 to the reference pattern of 16Sr group VI, subgroup A (Acc. No. AY390261). Here we report that the pear tree is an alternate host of the 'Ca. P. trifolii'.
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    Molecular phylogeny based on its sequences of nrDNA of some species belonging to dodder (Cuscuta L.) genus from various ecological sites of Turkey
    (Academic Press, 2020) Demir, İbrahim; Kaya, İlhan; Usta, Mustafa; Sipahioǧlu, Hikmet Murat
    Nuclear ribosomal DNA (nrDNA) sequence data of the Cuscuta genus, which have been considered as one of the most popular sequences for phylogenetic inference in plants, have been studied from a phylogenetic perspective in agricultural and non-agricultural lands of Turkey. The samples of Cuscuta spp. were collected from different geographical regions of Turkey between the years of 2013–2015. Some other species, not available locally, were taken from the herbarium samples of some research units. In order to study the phylogenetic relations of collected species, DNA isolations were made from body tissue samples. Conserved regions on ribosomal DNA (rDNA) were amplified by universal primers via PCR method and cloned into a proper cloning vector. The cloned DNA fragments were sequenced and analysed by web-based and computer programs. DNA sequences of certain species were recorded to the National Center for Biotechnology Information (NCBI) database. Based on the morphological examination and molecular analyses of fresh and the herbarium specimen, 8 species were identified. The identified species were C. hyalina (Gene bank accession no. KY020420), C. monogyna (KY020421), C. europaea (KY020422), C. palaestina (KY020423), C. approximata (KY020424), C. kurdica (KY020427), C. kotschyana (KY020430) and C. babylonica (KY020431). The ITS (Internal Transcribed Spacer) region contains several indels in identified Cuscuta species with the length varying from 668 to 730 bp. Sequence divergence ranges from 1.00% to 8.00% within Cuscuta spp. Based on our findings, the ITS sequences provided phylogenetically informative results in combination with the secondary structures.