Simplifying sample preparation by transferring plant sap (from plant sections) directly onto a me... more Simplifying sample preparation by transferring plant sap (from plant sections) directly onto a membrane is advantageous for any routine viroid detection technique, such as tissue printing. After fixing the samples on the membrane, hybridization steps similar to Northern or dot blot can be successfully implemented as long as factors like stringency and low viroid titer are properly adjusted to enable enhancement of the detection limit. The protocol described allows for indexing hundreds of field samples as a phytosanitary control measure.
Apple scar skin viroid (ASSVd) is a fruit-damaging pathogen that causes significant economic loss... more Apple scar skin viroid (ASSVd) is a fruit-damaging pathogen that causes significant economic losses to pome fruit trees. In the context of a survey on fruit tree viroids in Greece, ASSVd was initially detected by RT-PCR in two sweet cherry trees of cv. Tragana Edessis in an apple orchard in Florina (Macedonia, Greece). This finding was confirmed by direct viroid sequencing of the amplified RT-PCR products. In order to verify this finding, we further examined four sweet cherry trees cvs Tragana Edessis and Biggareau Burlat, two sweet cherry trees of undetermined cultivar, and fifteen neighboring apple trees in the same orchard for possible infection with ASSVd. The viroid assay was done by tissue print hybridization using an ASSVd-specific DIG-labeled probe at stringent hybridization conditions and by RT-PCR using two different ASSVd-specific primer pairs. ASSVd was detected in the six sweet cherry trees, including symptomatic samples, but not in any of the 15 apple trees. Purified A...
Symptoms of olive sickle leaf, a disorder of olive considered to be of virus or virus-like etiolo... more Symptoms of olive sickle leaf, a disorder of olive considered to be of virus or virus-like etiology, have been observed in Greece since 1980. Surveys at many places in Attiki, Peloponnisos and Makedonia regions showed symptoms in all cases, on cultivars Kalamon, Koroneiki, Manaki and Megaritiki. The frequency of symptom occurrence in individual trees and the intensity of symptoms varied from year to year, and 1981 was the year with the most frequent and strongest expression of the disorder. The proportion of affected shoots per tree usually varies between 5 and 90%.
A strain of artichoke Italian latent virus (AILV) was recently found infecting artichoke (Cynara ... more A strain of artichoke Italian latent virus (AILV) was recently found infecting artichoke (Cynara cardunculus v. scolymus L.) plants in the Argos area of Peloponnesus, in Greece (Rana and Kyriakopoulou, 1982). Infected plants were patchily distributed in the field and appeared yellowish and stunted (Fig. 1). Soil samples from the rhizosphere of these plants contained large numbers (500-600 individuals per 1/2 kg soil) of a dorylaimoid nematode described as a new species, Longidorus fasciatus, by Roca and Lamberti (1981). The ability of this nematode to transmit AILV was tested and the results of the transmission trials are reported in this paper.
Viroid research on pome and stone fruit trees in Greece is important, as it seems that such viroi... more Viroid research on pome and stone fruit trees in Greece is important, as it seems that such viroids are widespread in the country and may cause serious diseases. Our research dealt with three Pospiviroidae species infecting pome and stone fruit trees, namely Apple scar skin viroid (ASSVd), Pear blister canker viroid (PBCVd) and Hop stunt viroid (HSVd). Tissue-print hybridization, reverse transcription-polymerase chain reaction (RT-PCR), cloning and sequencing techniques were successfully used for the detection and identification of these viroids in a large number of pome and stone fruit tree samples from various areas of Greece (Peloponnesus, Macedonia, Thessaly, Attica and Crete). The 58 complete viroid sequences obtained (30 ASSVd, 16 PBCVd and 12 HSVd) were submitted to the GenBank. Our results showed the presence of ASSVd in apple, pear, wild apple (Malus sylvestris), wild pear (Pyrus amygdaliformis) and sweet cherry; HSVd in apricot, peach, plum, sweet cherry, bullace plum (Pru...
Apple scar skin viroid (ASSVd) and Pear blister canker viroid (PBCVd) are members of the genus Ap... more Apple scar skin viroid (ASSVd) and Pear blister canker viroid (PBCVd) are members of the genus Apscaviroid (family Pospiviroidae). In order to study the nucleotide sequence and secondary structure of Hellenic variants of these viroids, a large number of collected samples were initially screened by imprint hybridization; then ASSVd and PBCVd positive samples were assayed for the viroids by RT-PCR. Total RNA extracts were reverse-transcribed and amplified by polymerase chain reaction using two different specific primer pairs for each viroid. Purified RT-PCR products were directly sequenced or cloned into the pGEM-T and pCR® II vectors and then sequenced. Fourteen Hellenic full length ASSVd variants from 3 apple, 3 wild apple (Malus sylvestris), 1 wild pear (Pyrus amygdaliformis) and 3 pear trees are 330-335 nucleotides long. They differ from the reference sequences of ASSVd (ASSCS and Y00435) at 15-29 and 3-36 sites, respectively. Fifteen nucleotide changes (differences from ASSCS) ar...
... 23 PEACH LATENT MOSAIC VIROID IN TEMPERATE FRUIT HOSTS A. Hadidi, L. Giunchedi, H. Osaki, AM ... more ... 23 PEACH LATENT MOSAIC VIROID IN TEMPERATE FRUIT HOSTS A. Hadidi, L. Giunchedi, H. Osaki, AM Shamloul, A. Crescenzi, P. Gentit, L ... 2001). PLMVd from the above hosts may also be detected by RT-PCR-probe capture hybridization (ELISA) assay which was applied ...
Citriculture is one of the basic agricultural industries of Greece, with commercial production oc... more Citriculture is one of the basic agricultural industries of Greece, with commercial production occurring in 29 prefectures comprising about 25 million trees and producing 1.3 million tons of fruit, of which 40% is exported. Sweet orange is the dominant species, followed by lemon, mandarin and grapefruit, all grown exclusively on sour orange rootstock. Psorosis A was first observed in 1960, and is still found in old orchards, with the psorosis B form sometimes present. Concave gum is widespread, and impietratura, cristacortis, crinkly leaf, ringspot, exocortis, cachexia and gummy bark also occurring. Citrus tristeza virus was first detected in 2000 in a few illegally imported plants which, along with trees produced from them, have been destroyed. Measures to eradicate the virus continue. A new disease, sour orange woody gall, has also been found. The use of virus-tested budwood began in the early 1970s, and mother trees established with propagation material introduced from Corsica an...
Artichoke plants infected by raspberry ringspot virus (RRV) were found in Greece some years ago (... more Artichoke plants infected by raspberry ringspot virus (RRV) were found in Greece some years ago (Kyriakopoulou and Bern, 1982). RRV was isolated from plants showing yellow blotches on their leaves and from some without symptoms. Two artichoke strains of the virus, one from Greece and the other from Turkey and serologically different from the English and Scottish strains, were recently studied with regard to biological, physico-chemical and serological properties (Rana et al., 1985). RRV is transmitted in nature by tv,;o Longidorlis species: L. elongatus (de Man) Thorne et Swanger, (Taylor, 1962), and L. macrosoma Hooper (Harrison, 1964). Investigations on the distribution of RRV and its nematode vectors in artichoke fields in Greece were carried out during spring 1984 and the results from these are reported here.
Simplifying sample preparation by transferring plant sap (from plant sections) directly onto a me... more Simplifying sample preparation by transferring plant sap (from plant sections) directly onto a membrane is advantageous for any routine viroid detection technique, such as tissue printing. After fixing the samples on the membrane, hybridization steps similar to Northern or dot blot can be successfully implemented as long as factors like stringency and low viroid titer are properly adjusted to enable enhancement of the detection limit. The protocol described allows for indexing hundreds of field samples as a phytosanitary control measure.
Apple scar skin viroid (ASSVd) is a fruit-damaging pathogen that causes significant economic loss... more Apple scar skin viroid (ASSVd) is a fruit-damaging pathogen that causes significant economic losses to pome fruit trees. In the context of a survey on fruit tree viroids in Greece, ASSVd was initially detected by RT-PCR in two sweet cherry trees of cv. Tragana Edessis in an apple orchard in Florina (Macedonia, Greece). This finding was confirmed by direct viroid sequencing of the amplified RT-PCR products. In order to verify this finding, we further examined four sweet cherry trees cvs Tragana Edessis and Biggareau Burlat, two sweet cherry trees of undetermined cultivar, and fifteen neighboring apple trees in the same orchard for possible infection with ASSVd. The viroid assay was done by tissue print hybridization using an ASSVd-specific DIG-labeled probe at stringent hybridization conditions and by RT-PCR using two different ASSVd-specific primer pairs. ASSVd was detected in the six sweet cherry trees, including symptomatic samples, but not in any of the 15 apple trees. Purified A...
Symptoms of olive sickle leaf, a disorder of olive considered to be of virus or virus-like etiolo... more Symptoms of olive sickle leaf, a disorder of olive considered to be of virus or virus-like etiology, have been observed in Greece since 1980. Surveys at many places in Attiki, Peloponnisos and Makedonia regions showed symptoms in all cases, on cultivars Kalamon, Koroneiki, Manaki and Megaritiki. The frequency of symptom occurrence in individual trees and the intensity of symptoms varied from year to year, and 1981 was the year with the most frequent and strongest expression of the disorder. The proportion of affected shoots per tree usually varies between 5 and 90%.
A strain of artichoke Italian latent virus (AILV) was recently found infecting artichoke (Cynara ... more A strain of artichoke Italian latent virus (AILV) was recently found infecting artichoke (Cynara cardunculus v. scolymus L.) plants in the Argos area of Peloponnesus, in Greece (Rana and Kyriakopoulou, 1982). Infected plants were patchily distributed in the field and appeared yellowish and stunted (Fig. 1). Soil samples from the rhizosphere of these plants contained large numbers (500-600 individuals per 1/2 kg soil) of a dorylaimoid nematode described as a new species, Longidorus fasciatus, by Roca and Lamberti (1981). The ability of this nematode to transmit AILV was tested and the results of the transmission trials are reported in this paper.
Viroid research on pome and stone fruit trees in Greece is important, as it seems that such viroi... more Viroid research on pome and stone fruit trees in Greece is important, as it seems that such viroids are widespread in the country and may cause serious diseases. Our research dealt with three Pospiviroidae species infecting pome and stone fruit trees, namely Apple scar skin viroid (ASSVd), Pear blister canker viroid (PBCVd) and Hop stunt viroid (HSVd). Tissue-print hybridization, reverse transcription-polymerase chain reaction (RT-PCR), cloning and sequencing techniques were successfully used for the detection and identification of these viroids in a large number of pome and stone fruit tree samples from various areas of Greece (Peloponnesus, Macedonia, Thessaly, Attica and Crete). The 58 complete viroid sequences obtained (30 ASSVd, 16 PBCVd and 12 HSVd) were submitted to the GenBank. Our results showed the presence of ASSVd in apple, pear, wild apple (Malus sylvestris), wild pear (Pyrus amygdaliformis) and sweet cherry; HSVd in apricot, peach, plum, sweet cherry, bullace plum (Pru...
Apple scar skin viroid (ASSVd) and Pear blister canker viroid (PBCVd) are members of the genus Ap... more Apple scar skin viroid (ASSVd) and Pear blister canker viroid (PBCVd) are members of the genus Apscaviroid (family Pospiviroidae). In order to study the nucleotide sequence and secondary structure of Hellenic variants of these viroids, a large number of collected samples were initially screened by imprint hybridization; then ASSVd and PBCVd positive samples were assayed for the viroids by RT-PCR. Total RNA extracts were reverse-transcribed and amplified by polymerase chain reaction using two different specific primer pairs for each viroid. Purified RT-PCR products were directly sequenced or cloned into the pGEM-T and pCR® II vectors and then sequenced. Fourteen Hellenic full length ASSVd variants from 3 apple, 3 wild apple (Malus sylvestris), 1 wild pear (Pyrus amygdaliformis) and 3 pear trees are 330-335 nucleotides long. They differ from the reference sequences of ASSVd (ASSCS and Y00435) at 15-29 and 3-36 sites, respectively. Fifteen nucleotide changes (differences from ASSCS) ar...
... 23 PEACH LATENT MOSAIC VIROID IN TEMPERATE FRUIT HOSTS A. Hadidi, L. Giunchedi, H. Osaki, AM ... more ... 23 PEACH LATENT MOSAIC VIROID IN TEMPERATE FRUIT HOSTS A. Hadidi, L. Giunchedi, H. Osaki, AM Shamloul, A. Crescenzi, P. Gentit, L ... 2001). PLMVd from the above hosts may also be detected by RT-PCR-probe capture hybridization (ELISA) assay which was applied ...
Citriculture is one of the basic agricultural industries of Greece, with commercial production oc... more Citriculture is one of the basic agricultural industries of Greece, with commercial production occurring in 29 prefectures comprising about 25 million trees and producing 1.3 million tons of fruit, of which 40% is exported. Sweet orange is the dominant species, followed by lemon, mandarin and grapefruit, all grown exclusively on sour orange rootstock. Psorosis A was first observed in 1960, and is still found in old orchards, with the psorosis B form sometimes present. Concave gum is widespread, and impietratura, cristacortis, crinkly leaf, ringspot, exocortis, cachexia and gummy bark also occurring. Citrus tristeza virus was first detected in 2000 in a few illegally imported plants which, along with trees produced from them, have been destroyed. Measures to eradicate the virus continue. A new disease, sour orange woody gall, has also been found. The use of virus-tested budwood began in the early 1970s, and mother trees established with propagation material introduced from Corsica an...
Artichoke plants infected by raspberry ringspot virus (RRV) were found in Greece some years ago (... more Artichoke plants infected by raspberry ringspot virus (RRV) were found in Greece some years ago (Kyriakopoulou and Bern, 1982). RRV was isolated from plants showing yellow blotches on their leaves and from some without symptoms. Two artichoke strains of the virus, one from Greece and the other from Turkey and serologically different from the English and Scottish strains, were recently studied with regard to biological, physico-chemical and serological properties (Rana et al., 1985). RRV is transmitted in nature by tv,;o Longidorlis species: L. elongatus (de Man) Thorne et Swanger, (Taylor, 1962), and L. macrosoma Hooper (Harrison, 1964). Investigations on the distribution of RRV and its nematode vectors in artichoke fields in Greece were carried out during spring 1984 and the results from these are reported here.
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Papers by P. Kyriakopoulou