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    Diyarbakır ili domates üretim alanlarındaki bazı önemli virüslerin moleküler yöntemler ile teşhisi ve moleküler karakterizasyonu
    (Zirai Mücadele Merkez Araştırma Enstitüsü, 2020) Yılmaz, Feyzullah; Sipahioğlu, Hikmet Murat
    Diyarbakır ili domates üretim alanlarında Domates sarı yaprak kıvırcıklık virüsü (Tomato yellow leaf curl virus, TYLCV), Patates Y virüsü (Potato virus Y, PVY), Domates mozaik virüsü (Tomato mosaic virus, ToMV), Domates lekeli solgunluk virüsü (Tomato spotted wilt virus, TSWV) ve Domates halkalı leke virüsü (Tomato ringspot virus, ToRSV)’nün varlığının moleküler yöntemler ile ortaya konulması amacıyla 2018 yılında sürvey çalışmaları yürütülmüştür. TYLCV’nin varlığını araştırmak için PCR yöntemi, PVY ve ToMV’nin varlığını araştırmak için multipleks RT-PCR yöntemi, TSWV ve ToRSV’nin varlığını araştırmak için ise RT-PCR yöntemi kullanılmıştır. Domates üretim alanlarında virüs belirtisi gösteren ve göstermeyen toplam 278 adet bitkiden rastgele yaprak örnekleri toplanmıştır. Moleküler yöntemler ile testlenen örneklerin 56 (%20.1)’sının en az bir virüs türü ile enfekteli olduğu tespit edilmiştir. Testlenen örneklerin 41’inin (%14.7) ToMV, 21’nin (%7.6) PVY ve 6’sının (%2.2) ise ToMV+PVY ile karışık enfekteli olduğu saptanmıştır. Çalışma kapsamında yürütülen moleküler testler sonucunda, domates örneklerinde TYLCV, TSWV ve ToRSV enfeksiyonları saptanmamıştır. Tespit edilen PVY ve ToMV izolatlarından rastgele 4’er adet seçilerek PVY izolatlarının kısmi kılıf protein genleri, ToMV izolatlarının kısmı hareket protein ve kılıf protein genleri karakterize edilerek, Gen Bankasına kayıtları yapılmıştır. ToMV izolatları dünyadaki diğer izolatlar ile nükleotid düzeyinde %99-100 arasında benzerlik gösterirken, PVY izolatları %88-99 arasında değişen oranda benzerlik göstermiştir. Yürütülen bu çalışma ile ToMV ve PVY Diyarbakır ili domates üretim alanlarında ilk defa rapor edilmiştir.
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    First Detection and Molecular Characterization of Peach latent mosaic viroid in a New Natural Host: Walnut (Juglans regia L.)
    (2020) Tuncel, Feyza; Tekkaş, Nazlı; Türk, Gökay; Oksal, Hatice Diğdem; Sipahioğlu, Hikmet Murat
    Peach latent mosaic viroid (PLMVd) occurs naturally in peach and nectarines worldwide including North and South America, Europe, Asia and Australia with high infection frequencies. Here, our experimental findings revealed for the first time PLMVd infections in walnut (Juglans regia L.) in the world. Twelve walnut leaves, all from symptomless plants, were obtained from a nearby walnut germplasm collection in Malatya (Turkey). Genomic RNAs were extracted using a commercial RNA isolation kit and the complete viroid genome was amplified by reverse transcriptase polymerase chain reaction (RT-PCR) assay using end to end primer pair. The PCR amplicons of expected size (approx. 0.34-kb DNA fragments) were cloned into pGEM-T Easy vector and transformed into competent Escherichia coli strain JM109. Purified plasmids were sequenced bidirectionally. Of the 12 walnut specimens tested by RT-PCR, 4 were found infected by PLMVd detected in mid-summer of 2018 and in autumn of 2019. Analysis of nucleotide sequence of two walnut isolates (Acces. no: MN857143- MN857144) confirmed the presence of PLMVd and shared 96.6-98.8% similarity (Acces. no: MN857143) and %97-98.8% similarity (Acces. no: MN857144) with peach, nectarine and cherry isolates of the world. By identifying walnut as an alternate host, we confirm that PLMVd potentially may infect other fruit trees cultivated worldwide. Here, we also report molecular features of these two distinct PLMVd-walnut isolates detected in Malatya (Turkey).
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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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    Occurrence and Molecular Characterization of Some Economically Relevant Cucurbit Viruses in Malatya, Turkey
    (Adem Yavuz Sönmez, 2020) Örs, Fatih; Oksal, Hatice Diğdem; Sipahioğlu, Hikmet Murat
    Field surveys were conducted from 2016 to 2017 to detect and determine the prevalence of viruses in the major cucurbit growing areas of Malatya. A total of 162, namely 110 melon, 25 pumpkin and 27 watermelon leaf samples were collected from Arapgir, Arguvan and Battalgazi districts from symptomatic and non-symptomatic cucurbit plants. Using molecular methods, the plant samples were tested against four viruses, including Cucumber mosaic cucumovirus (CMV), Zucchini yellow mosaic potyvirus (ZYMV), Watermelon mosaic potyvirus (WMV) and Squash mosaic comovirus (SqMV). Results of RT-PCR tests revealed that none of the samples collected from Arapgir and Arguvan reacted positive against surveyed viruses. However, CMV infection was found in 1 cucumber, ZYMV in 5 pumpkin and WMV in 6 watermelon samples in Battalgazi district. The nucleotide sequences of partial coat protein (CP) genes of ZYMV-Malatya and WMV-Malatya isolates were determined. Nucleotide sequence comparisons revealed 95-100% similarity of ZYMV-Malatya isolate with other isolates of ZYMV reported worldwide with no mutation site on CP gene. However, WMV-Malatya isolate exhibited 94-99% similarity with other WMV sequences reported worldwide consisting a unique mutation site on CP gene sequence. These isolates are recorded to the Gene Bank with the accession numbers MT186266 (Zucchini yellow mosaic virus isolate, Malatya ZYMV1) and MT186267 (Watermelon mosaic virus isolate Malatya WMV1).