Browsing by Author "Guller, Abdullah"

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Antiviral and Antifungal Activity of Biologically Active Recombinant Bouganin Protein From Bougainvillea Spectabilis Willd
(Ankara Univ, Fac Agr, 2018) Guller, Abdullah; Siphioglu, Hikmet Murat; Usta, Mustafa; Demirer Durak, Emre
Bouganin antiviral protein (BAP) gene. one of the ribosome inactivating proteins, isolated from Bougainvillea speclabilis Willd. was cloned, expressed and the antiviral and antifungal activities were investigated. The full-length bouganin antiviral protein gene was amplified by reverse transcription-PCR using mRNA as template extracted from mature leaves. The coding region of bouganin gene was cloned into prokaryotic expression vector pETDuet-1 after amplification with end to end gene specific primers. The recombinant plasmid was transformed into Escherichia coli cells BL21(DE3)pLysS and the expression of BAP gene was induced by isopropyl beta-D thiogalactopyranoside (IPTG). Bouganin antiviral protein having a molecular mass of 28 kDa has been isolated from transformed bacterial colonies. Antiviral activity of bouganin was assayed against Zucchini yellow mosaic virus (ZYMV) by a mechanical inoculation test. The antifungal activity of purified recombinant protein was tested against pathogenic and non-pathogenic Rhizocionia solani. Trichoderma harzianum, and Fusarium arysporurn fungi using disc diffusion method. The increased amount of antiviral protein reduced the disease severity caused by ZYMV. The bouganin antiviral protein was inhibited the growth of R. solcati by 30.7% and of T harzianum by 20% after 72 h compared to control. No growth inhibition was observed for F oayporurn. All plants including controls treated with in vitro expressed BAP protein exhibited severe growth reduction compared with negative control (not treated) plants.
Coat Protein of Alfalfa Mosaic Alfamovirus (Amv) From Türkiye: Genetic Inference and in Silico Docking Analysis for Potential Antiphytoviral Purposes
(Univ Agr Sci & veterinary Med Cluj-napoca, 2024) Demirel, Serap; Guller, Abdullah; Usta, Mustafa; Kurt, Zeynelabidin; Korkmaz, Gulustan
In 2021, a study was conducted in the Denizli region of Turkiye to investigate the phylogenetic relationship and presence of alfalfa mosaic alfamovirus (AMV) infecting pepper plants exhibiting viral disease symptoms. A total of 57 samples were collected, of which twenty-four tested positive for AMV with reverse transcriptase polymerase chain reaction (RT-PCR) assays. Samples from pepper plants displaying virus symptoms gave positive bands on the agarose gel, while healthy plants yielded negative results. One of the positive samples was randomly selected, cloned and sequenced. This sequence of the Denizli AMV isolate (753 bp) was recorded in the GenBank database with accession number OQ845956. The nucleotide sequence showed a high nucleotide consensus of 97%-99% compared with the nucleotide sequences of the same variants from different origins in GenBank. According to the phylogenetic tree generated through the Neighbour Joining (NJ) method, this AMV isolate belongs to the same group as Iranian isolates from various of hosts. Furthermore, in silico docking analysis of the coat protein (CP) of the AMV isolate and promising 12 essential oil compounds was performed to enable potential antiviral drug development. Docking study showed that eucalyptol, eugenol and carvacrol can make important contributions to the advancement of drug -based strategies for the managing of plant viruses by interacting with the virus coat protein of high binding energies, -5.3, -5.2 and -5.0 kcal mol-1, respectively. Although the presence of AMV in Denizli province has been reported previously, this study reports the phylogenetic relationships and docking analysis of the new AMV isolate in pepper crops.
Comprehensive Survey of Common Potato Viruses in Eastern Anatolia Region of Turkey: New Isolates and Phylogenetic Insights
(Springer, 2025) Korkmaz, Gueluestan; Usta, Mustafa; Guller, Abdullah; Demirel, Serap
Potatoes (Solanumtuberosum L.) are a vital crop in Turkey, contributing significantly to the country's economy. However, they are susceptible to various viral infections, causing substantial yield losses if left unmanaged. This study aimed to conduct a comprehensive survey of economically significant viruses affecting potatoes in the Eastern Anatolia Region, Turkey, including the identification of new isolates and the inference of their phylogenetic relationships. A total of 1130 fresh leaf samples, including asymptomatic and symptomatic plant samples, were tested using the multiplex RT-PCR (m-RT-PCR) method to determine the prevalence of five viral pathogens. The test results revealed the presence of four viral pathogens, with an overall infection incidence of 37.08%. PVY was found in 35% of the samples, PVS in 8.5%, PVX in 1.8%, and PLRV in 1%. However, PVA was not detected. In addition, 74 of the 98 samples tested were positive for PVY + PVS, 13 for PVY + PVX, 2 for PVY + PLRV, 8 for PVY + PVS + PVX, and 1 for PVY + PVS + PLRV. From each sample that tested positive, two isolates were selected, and the complete coat protein (CP) gene regions of these isolates were cloned and sequenced. The sequences of the new potato virus isolates were deposited in the GenBank. Nucleic acid sequence comparisons revealed a consensus of over 99% among all available isolates and those from other regions of the world. However, this similarity is not specific to any particular host or region. In support of the similarities in nucleic acid, phylogenetic analyses also provided information on the evolutionary relatedness of these isolates from various origins and hosts. This finding suggests that the genetic makeup of these isolates is highly conserved across different populations and environments. This study, conducted in the Eastern Anatolia Region of Turkey, is the first comprehensive report on the presence and molecular phylogeny of four viral pathogens of economic importance in potato crops.
Current Status and Molecular Phylogeny of Some Economically Important Viruses Infecting Wheat Crops of Sirnak Province, Turkey
(United Arab Emirates Univ, 2023) Kapan, Orhan; Usta, Mustafa; Guller, Abdullah
A survey of yellow dwarf viruses (YDVs)-associated diseases of wheat plants was performed for the first time in the Sirnak Province, Turkey. In plants wheat cultivated areas, symptoms such as diminished leaf size, dwarfing, yellowing, and reddening of leaves wheat plants were observed in cultivated areas. A total of 441 specimens were collected regardless of showing symptoms and assayed for Barley/Cereal Yellow Dwarf Viruses [B/CYDV] association by PCR tests using virus coat protein gene (CPG)-specific primer pairs. These assays demonstrated that 13.38% (55 samples) were positive to the occurrence of BYDV-PAV (9.73%) and CYDV-RPV (0.73%) for wheat viruses, but no BYDV-MAV,-SGV, and-RMV infections of BYDVs were found in PCR assays of collected samples. In addition, double infection of BYDV-PAV and CYDV-RPV was detected in 12 samples with a 2.91% infection rate. The CPG sequences of randomly selected two isolates were revealed using bacterial cloning and sequencing. Sequences obtained were deposited in GenBank under Accession numbers OL685734 and OL685736 for BYDV-PAV; and OL685735 and OL685737 for CYDV-RPV. BLASTn analysis of CPG sequences of four isolates shared high nucleotide homology with their intraspecies isolates, with 97.35-98.67% for two BYDV-PAV and 97.72-97.89 for two CYDV-RPV. The consensus tree constructed classified B/CYDV-associated wheat viruses with their phylogenetically closest individuals from Turkey and the world. This survey is the first report of BYDV-PAV and CYDV-RPV associated with wheat plants in Sirnak Province of Turkey.
Detection, in Silico Analysis and Molecular Diversity of Phytoplasmas From Solanaceous Crops in Turkey
(Czech Academy Agricultural Sciences, 2022) Usta, Mustafa; Guller, Abdullah; Sipahioglu, Hikmet Murat
Phytoplasma-like symptoms of leaf yellowing and calyx malformation were observed in eggplant (Solanum melongena L.), upward leaves and fruit malformation in pepper (Capsicum annuum L.), and aerial tuber formation in potato (S. tuberosum L.) during the survey performed in the late season (August to September) of 2015 and 2016 in Van province (Turkey). A total of 100 samples were tested by nested-PCR using universal primer pairs to assess the sanitary status of the solanaceous crops and to characterise the phytoplasma isolates. Among them, seven sam-ples resulted in a 1.25 kb DNA fragment, and five (two eggplants, two peppers, and one potato) were molecularly characterised (Accession No.: KY579357, KT595210, MF564267, MF564266, and MH683601). BLAST and the virtual restriction fragment length polymorphism (RFLP) analysis of 16S rRNA genes revealed the presence of two distinct phytoplasma infections in solanaceous crops: `Candidatus Phytoplasma trifolii' a member of the clover proliferation group (16SrVI) and subgroup A and `Candidatus P. solani' a member of the stolbur group (16SrXII) and subgroup A. The virtual RFLP analysis and calculated coefficients of RFLP pattern similarities further revealed a remarkable ge-netic diversity among the `Candidatus P. solani' isolates infecting pepper (similarity coefficient of 0.90) and eggplant (similarity coefficients of 0.98 and 1.00) at the same geographical area. This is the first report of the natural occurrence of `Candidadtus P. trifolii' in potato from the Eastern Anatolia region, Turkey.
First Report of "candidatus Phytoplasma Solani" on a New Host Marigold (Tagetes Erecta L.)
(Tubitak Scientific & Technological Research Council Turkey, 2016) Alp, Sevket; Usta, Mustafa; Sipahioglu, Hikmet Murat; Guller, Abdullah
Marigold (Tagetes erecta L.) plants, also called Mexican or Aztec marigold, with symptoms of shoot proliferation, dwarfing, and reddening were observed in ornamental gardens of Van Province (Turkey). Five plants, two of them showing reddening and three symptomless plants, were sampled at the end of September 2014. Genomic DNA isolated from symptomatic and nonsymptomatic plant leaves was used to amplify 16S rDNA fragments by nested polymerase chain reaction (PCR). Of the 5 marigold samples tested by PCR, only the two showing reddening symptoms yielded the expected 1.2-kb DNA fragments. Amplified PCR fragments were cloned into a plasmid vector and transformed into competent Escherichia coli strain JM 109. Recombinant plasmid DNA was isolated and sequenced bidirectionally. The provided sequences were 1244 bp and 1245 bp in length and were designated as isolate 1 and isolate 2, respectively. BLAST analysis of the 16S rDNA sequence and virtual restriction fragment length polymorphism (RFLP) analysis confirmed the presence of the phytoplasma "Candidatus Phytoplasma solani". The in silico virtual RFLP pattern of isolate 1, based on the 16S rDNA F2n/R2 fragment, was identical (similarity coefficient 1.00) to the reference pattern of 16Sr group XII, subgroup A (GenBank accession no.: AF248959). Isolate 1 was identified as a member of 16SrXII-A. Based on the same analyses, isolate 2 showed molecular characteristics different from reference patterns of all previously established 16Sr groups and subgroups. The most similar was the reference pattern of 16Sr group XII, subgroup A (GenBank accession no.: AF248959), with a similarity coefficient of 0.97. This is the first report of naturally occurring "Ca. P. solani" affecting T. erecta, which shows that this plant species is an alternate host of the agent.
First Report of 'candidatus Phytoplasma Trifolii' Associated With Leaf Reddening and Upright Growth in Pears (Pyrus Communis L.)
(Czech Academy Agricultural Sciences, 2021) Usta, Mustafa; Guller, Abdullah; Sipahioglu, 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 midvein, 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 sequences, 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'.
First Report of Natural Infection of Watermelon Mosaic Virus (Wmv) Infecting Bottle Gourd and Snake Melon
(Univ Namik Kemal, 2024) Guller, Abdullah; Usta, Mustafa; Korkmaz, Gulustan; Demirel, Serap
Cucurbitaceous crops, one of the main crops of agriculture, are sensitive to many plant viruses. In August 2019, virus-like symptoms were observed on some cucurbit plants grown in private home gardens in Antalya and Denizli provinces (Turkey). A total of 53 leaf samples were sampled from plants with the most symptoms (melon (Cucumis melo L.), watermelon (Citrullus lanatus L.), bottle gourd (Lagenaria siceraria (Molina) Standl.), and snake melon (Cucumis melo var. flexuosus) and tested by Reverse-Transcriptase Polymerase Chain Reaction (RT-PCR) against possible watermelon mosaic potyvirus (WMV) infection. The coat protein gene (CP) specific primer sets amplified a gene product of nearly 820 bp fragment from symptomatic plants. WMV infections were detected in 31 individual cucurbit plants, including 11 melons, 8 watermelons, 7 snake melons and 5 bottle gourds. The presence of viral infection was found only in ornamental squash plants in Antalya province and in all cucurbits sampled in Denizli province. To better comprehend the molecular characteristics of virus isolates, the amplified viral DNA fragments were cloned in a proper prokaryotic plasmid, sequenced by Next Generation Sequencing (NGS) and recorded to GenBank. Bioinformatic analyses using the Basic Local Alignment Search Tool (BLAST) showed that the identified CP gene sequences exhibited significant nucleotide homogeneity, supported by a high nucleotide similarity index with that of other isolates around the world. In addition, Turkish isolates isolated from Antalya and Denizli regions showed approximately 94% nucleotide similarity among themselves. For phylogenetic inference, WMV sequences were subjected to multiple alignments with isolates from different geographic origins of the same viruses. Molecular phylogeny showed that all WMV isolates are closely related to other world WMV isolates at variable rates. WMV is wide host range viruses in cucurbit crops, however, this work is the first scientific report of WMV isolates detected in bottle gourd and snake melon from the South and West Regions of Turkey all over the world.
Incidence and Molecular Characterisation of Viruses Infecting Wheat in Eastern Anatolia (Turey)
(Parlar Scientific Publications (p S P), 2020) Usta, Mustafa; Sipahioglu, H. Murat; Guller, Abdullah
In order to assess the occurrence and the incidence of Barley yellow dwarf viruses (BYDV-PAV, BYDV-SGV, CYDV-RPV, BYDV-MAV, BYDV-RMV), Wheat streak mosaic virus (WSMV) and Soil borne wheat mosaic virus (SBWMV) in eastern Anatolia, a survey was conducted in wheat fields from 10 major wheat-growing provinces in Turkey from April 2012 to June 2013. A total of 900 wheat samples were collected and reverse-transcription polymerase chain reaction (RT-PCR) and multiplex RTPCR tests were implemented to detect seven wheat viruses. In wheat fields, BYDV-PAV was the most common. followed by BYDV-SGV, CYDV-RPV and WSMV, respectively. BYDV-MAV, BYDV-RMV, and SBWMV were not detected in tested samples. Since the SBWMV has been transmitted by soil borne fungus Polymyxa graminis, further research was performed to determine the presence of SBWMV and its fungus vector in soil samples by bait plant method, None of the green parts and roots of the bait plants were found infected by SBWMV and P. gratninis, respectively. In the wheat fields of eastern Anatolia, total virus incidence ranged from 0,4 to 6.8%. Based on field observations, the incidence of surveyed viruses in all wheat fields was less than 7 %. Since detected viruses have been considered the most economically relevant viruses of wheat fields, we further characterized randomly selected isolate of each detected virus by cloning the complete coat protein gene and sequencing [(Accession numbers; BYDV-PAV (KC900900), CYDV-RPV (KC900903), BYDV-SGV (partial) (KC900899) and WSMV (KC900901)]. Nucleotide sequence identities for the coat protein gene (CP) ranged from 82 % to 94 % (PAV), 69 % to 96% (SGV), 83% to 96 % (CYDV-RPV), and from 88 0 to 97 % (WSMV). This is the first report of BYDV-PAV, BYDV-SGV, CYDV-RPV and WSMV infecting wheats in eastern Anatolia (Turkey).
Molecular Analysis of 'candidatus Phytoplasma Trifolii' and 'candidatus Phytoplasma Solani' Associated With Phytoplasma Diseases of Tomato (Pdt) in Turkey
(Friends Science Publ, 2018) Usta, Mustafa; Guller, Abdullah; Sipahioglu, Hikmet Murat
Tomato plants displaying severe fruit deformation, flower sterility, aerial rooting, purplish leaves and leaf rolling were observed in tomato fields at Van province (Turkey). Samples were collected, and total DNA was extracted from symptomatic and asymptomatic plants. Nested polymerase chain reaction (nested-PCR) assays were performed to amplify 16S rDNA sequences for molecular detection using universal primer pairs. Out of 100 tested tomato samples, 11% of tomato samples yielded a DNA fragment of 1.25 kb. Amplified PCR products were then cloned into pGEM T-Easy vector and sequenced using new generation DNA sequencing (NGS) system. The virtual restriction fragment length polymorphism (RFLP) analysis of 16S rDNA sequences and molecular detections were allowed to characterize possible phytoplasmas associated with diseased plants. Our results revealed the presence of two Phytoplasma species belonging to two different ribosomal groups; 'Candidatus Phytoplasma trifolii' (16Sr VI-A group) (Acces no. MF564268, MG732925) and 'Candidatus Phytoplasma solani' (16SrXII-A group) (Acces no. KY579358, MF576263). Despite a high variation in their similarity coefficient of'Ca. P. solani' VTS2 (0.91) and 'Ca. P. trifolii' VTT1 (0.88) isolates, the infected tomato plants generally displayed similar disease symptoms during field observations. Due to its commercial interest, co-existing of these phytoplasmas in tomato fields is of great phytosanitary significance not only for tomato plants but also for other crops such as vegetables, ornamentals and field crops. With this study, 'Ca. P. trifolii' associated with phytoplasma diseases of tomato (PDT) has been reported for the first time in tomato in Turkey. (C) 2018 Friends Science Publishers
The Molecular Characterization of the Coat Protein Sequence and Differentiation of Cmv- Subgroup I on Tobacco From Native Flora in Turkey
(Univ Agr Sci & veterinary Med Cluj-napoca, 2020) Usta, Mustafa; Guller, Abdullah; Gunay, Abidin
Cucumber mosaic virus (CMV) has a broad plant-host range and a wide ecological zone distribution. Virus-like symptoms were observed on tobacco fields of Adiyaman province (Turkey) showing conspicuous mottling, greenish mosaic patterns and severe malformations of leaves. A total of forty tobacco samples tested positive against CMV by reverse transcription polymerase chain reaction (RT-PCR) using coat protein gene specific primers. Five randomly chosen CMV isolates were cloned into pGEM T-Easy vector and transformed into Escherichia coli JM109 strain. The recombinant bacterial clones containing insert-DNA were further purified and sequenced bidirectionally. In multiplex-RT-PCR studies carried out, it was found that all 40 CMV isolates belong to Subgroup I by resulting a 593 bp long DNA fragments. CMV subgroup IA was found to predominate in 4 out of 5 tobacco samples and CMV subgroup IB was found in 1 out of 5 CMV-positive samples by comparing the isolates with CMV reference isolates in phylogenetic tree. However, no Subgroup II sequences were found by multiplex RT-PCR using discriminating primers. The nucleic acid sequences were analyzed for the investigation of diversity of coat protein (CP) sequences of 5 CMV isolates. The sequence similarity ranged from 94.2-100% with the CMV subgroup I isolates infecting diverse plants in other regions of the world. The evolutionary tree revealed that the CMV IA Adiyaman isolates exhibited a genetic affinity with Australian and Spanish isolates. However, the CMV IB Adiyaman isolate showed a close genetic relationship with only the Australian isolates. To our knowledge, this study shows for the first time the occurrence of CMV IA and IB isolates infecting cultured tobacco plants in Adiyaman province.