Journal of Plant Pathology (2013), 95 (3), 579-586 Edizioni ETS Pisa, 2013 Caglayan et al. 579

EVALUATION OF THE SUSCEPTIBILITY OF DIFFERENT ROOTSTOCKS TO NATURAL INFECTION OF VIRUS-T

K. Caglayan1, C.U. Serce1*, M. Gazel1, K. Kaya1, F.C. Cengiz1, E. Vidal2 and M. Cambra2

1Mustafa Kemal University, Faculty of Agriculture, Plant Protection Department, 31034 Hatay, Turkey 2Laboratorio de Virología e Inmunología, Centro de Protección Vegetal Biotecnología, Instituto Valenciano de Investigaciones Agrarias (IVIA), Ctra. Moncada-Náquera km 5, 46113 Moncada, Valencia *Present address: Nigde University, Faculty of Agricultural Science and Technologies, Department of Crop Production and Technologies 51240 Nigde, Turkey

SUMMARY INTRODUCTION

Plum pox virus (PPV) has been observed in Turkey In Turkey, Plum pox virus (PPV) was first recorded in since 1968, but was not widespread except in apricot the Edirne (Thrace) (Sahtiyancı, 1969) and subsequently and plum trees in home gardens and ornamental parks in Ankara (Central Anatolia) (Kurçman, 1973) provinces. in restricted areas. Susceptibility of six different Prunus More recent surveys revealed new PPV outbreaks in the rootstocks to strain PPV-T was assessed under natural Aegean and Mediterranean regions (Gümüş et al., 2007; inoculum pressure in the Izmir-Aegean region during Gazel et al., 2010). 2010-2011. populations were monitored from the There is a wide genetic variability within PPV (Can- first week of April to the middle of June by the sticky- dresse and Cambra, 2006; Barba et al., 2011). Several plant method one year after the rootstock plantation was strains or subgroups are recognized their classification established. collected from different rootstocks being based on biology, serology and molecular proper- were tested individually by squash real-time RT-PCR ties. The two most common strains are PPV-D (Dideron) and all rootstocks were regularly tested by DASI-ELISA. and PPV-M (Marcus). Additionally, five other strains have The largest aphid populations were observed at the end been characterized, i.e.. PPV-Rec (Recombinant), PPV-EA of May and the most abundant aphid species as averages (El Amar), PPV-W (Winona), PPV-C (Cherry) (Glasa et al., over the two years were Myzus persicae (20.15%), Hyalop- 2004b; James and Varga, 2005; Candresse and Cambra, terus pruni (18.64%), Aphis craccivora (9.04%) and Aphis 2006) and more recently, PPV-T (Turkey) (Ulubas Serce et gossypii (8.36%). In 2011, the highest percentage of virulif- al., 2009). Numerous PPV isolates were described as hav- erous aphids was found in M. persicae (34.78%), followed ing different biological and epidemiological characteristics, by H. pruni (32.50%), Macrosiphum euphorbiae (25.00%), such as those related to aggressiveness (Quiot et al., 1995), A. gossypii (23.80%), A. spiraecola (12.50%) and A. crac- aphid transmissibility (Deborré et al., 1995) and symptom- civora (10.00%). Of the six Prunus rootstocks tested, only atology (Jarausch et al., 2004; Palmisano et al., 2010). Se- Nemaguard and Myrobalan 29C were infected by PPV-T, rological characterization studies of Turkish PPV isolates infection rate in 2010 being 6.0% (Nemaguard) and 4.0% using universal and strain-specific monoclonal antibodies (Myrobalan 29C). The infection rate increased to 16.0% (MAbs) showed that fifteen PPV-positive apricot samples for Nemaguard and 14.0% for Myrobalan 29C in 2011. from different regions of Turkey reacted with MAb4DG5 However, the other rootstocks, Prunus marianna GF8.1, and MAbAL which made their serotyping difficult (Myrta Docera6, GF677 and Garnem tested negative for PPV-T et al., 1998). throughout 2011. PPV isolates obtained from naturally in- A similarly ambiguous result (presence of both M- and fected apricot trees (inoculum source) and from infected D- specific epitopes in the same isolate) was reported by rootstocks in the experimental plot were characterized Candresse et al. (1998) with a Turkish PPV isolate. Partial as PPV-T and had more than 99.5% nucleotide sequence sequencing of this isolate (Ab-Tk) revealed that it is a identity. novel type of recombinant, characterized by a recombi- nation breakpoint in the HC-Pro gene, around position 1,566 of the genome (Glasa and Candresse, 2005). Fur- Key words: Sharka disease, PPV Turkey strain, viru- ther studies showed that Ab-Tk and several other Turk- liferous aphids, rootstock susceptibility, epidemiology, ish PPV isolates should be considered as members of a detection. new PPV strain for which the name PPV-T was proposed (Ulubas Serce et al., 2009). Three PPV strains (PPV-M, Corresponding author: K. Caglayan PPV-Rec and PPV-T) have been reported in Turkey each Fax: +90.326.2455832 with a distinct distribution (Çağlayan et al., 2012). PPV-T, E-mail: [email protected]

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Fig. 1. Vein clearing and chlorosis on the leaves of Nemaguard (left) and Myrobalan 29C (right) rootstocks in the experimental plot due to Plum pox virus (T strain) infection.

originally detected and characterized in Turkey, prevails to standard nursery practices without any phytosanitary in central Anatolia (Ankara) and the Aegean regions treatment. (Izmir) where PPV has been endemic for years. PPV-M was reported primarily as new outbreaks in the eastern Monitoring of PPV spread. Serological tests were car- Mediterranean region and PPV-Rec was found only in ried out one year after rootstocks planting, i.e. April of Isparta, in the western Mediterranean region (Candresse 2010 and 2011. Plants were regularly checked for PPV et al., 2007). Although many studies are available on the symptoms and individually sampled by collecting four epidemiological aspect of different PPV-M and D isolates fully expanded leaves from different parts of the canopy. from different countries, no such data are available for Serological assay for PPV detection was by DAS-ELISA PPV-T. Therefore, in this study, the natural transmission based on the 5B-IVIA (Cambra et al., 1994) monoclonal status and rootstock reactions to PPV-T in the Aegean antibody kit (Plant Print Diagnostics, Spain), following the region of Turkey were investigated. EPPO (2004) protocol for PPV detection.

Molecular characterization of PPV isolates. Samples MATERIALS AND METHODS from nine PPV-positive rootstocks (six Myrobalan 29C, three Nemaguard) in the experimental plot and six apri- Plant material and experimental nursery plots. An cot samples randomly selected in the adjacent infected experimental nursery plot was established in a naturally orchard were analyzed for PPV strain identification in the PPV-infected apricot orchard in spring 2009 at İzmir (Ae- second year of evaluation. Leaves exhibiting PPV symp- gean region). Spread of PPV and incidence of major aphid toms were collected and RNA was extracted using a LiCl species were followed for two consecutive years, 2010 and method (Spiegel et al., 1996). Four μl of RNA was used 2011. for cDNA synthesis using a reverse transcription kit (MBI The susceptibility to natural PPV-T infection was Fermentas, Finland). PCR was performed using PPV uni- evaluated of six different in vitro-propagated one-year- versal primer pairs that amplify 745 bp fragments: NCU- old PPV-free certified Prunus rootstocks (Agromillora niFor 5’-GAGGCAATTTGTGCTTCAATGG-3’ and Iberia, Spain), i.e. Myrobolan 29C (P. cerasifera), Garnem NCUniRev 5’-CGCTTAACTCCTTCATACCAAG-3’ (P. dulcis × [(P. persica × Prunus davidiana) × P. persica]), (Predajna et al., 2012). Cycling parameters were: 95°C for P. marianna GF 8.1 (P. cerasifera × Prunus munsoniana), 3 min, followed by 35 cycles of 94°C for 30 sec, 53°C for Nemaguard (P. persica × P. davidiana), Docera 6 (P. do- 1 min and 72°C for 1 min, followed by 72°C for 10 min. mestica x P. cerasifera) and GF677 (Prunus amygdalus × P. PCR amplicons were sequenced directly and analyzed persica). Fifty plants for each rootstock were planted adja- using the basic local alignment search tool (BLAST). cent to a PPV-infected apricot orchard but some of them Multiple alignments of the nucleotide sequences of these died in the first year (Table 1). The plants were planted in isolates and reference isolates for each PPV strain ob- two rows parallel to the inoculum source by using longi- tained from GenBank were performed using BioEdit tudinal design of 5 groups or replicates of 10 plants into (Hall, 1999). Cluster analysis was done with the Mega 5.0 each group (20 cm apart). The groups were randomly dis- program (Tamura et al., 2011) using the neighbor-joining tributed. The experimental field was managed according method with nucleotide identity distances. Bootstrap

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Table 1. Natural incidence of Plum pox virus-T in different Prunus rootstocks determined by DAS-ELISA in an experimental nursery plot in Izmir/ Turkey.

Number of infected / tested plants Natural infection rate (%) Name of the rootstocks 2010 2011 2010 2011 Nemaguard 3/50 8/50 6 16 Myrobalan 29C 2/50 7/50 4 14 Garnem 0/47 0/47 0 0 P. marianna GF8.1 0/48 0/48 0 0 GF-677 0/50 0/50 0 0 Docera 6 0/40 0/40 0 0

Total plants 5/285 15/285 1.75 10.53

analyses with 1000 replicates were performed to estimate Characterization of PPV isolates. The sequences of the support for inferred phylogenies. PCR amplicons obtained from nine viral isolates recovered from the experimental plot were analyzed, showing 98% Monitoring of aphid species and detection of viru- identity with the AbTk isolate of PPV-T, 95% identity with liferous aphids. Adult winged aphid populations which the BOR-3 and Niksic6 isolates of PPV-Rec, 94% identity were visiting the rootstock plants in the experimental plot with the N1, SK68 and PS of PPV-M strains (GenBank ac- were monitored by the sticky-plant method (Avinent et al., cession Nos. EU734794, JQ794541, HQ452359, FJ361234, 1993; Marroquin et al., 2004) during spring 2010 and 2011. M92280 and AJ243957, respectively). The nucleotide se- Three sticky shoots from each rootstock species were col- quence identity of the nine above PPV isolates and of six lected each week and new shoots were sprayed for next additional isolates from the adjacent apricot orchard was collections. Collected shoots were processed to identify greater than 99.5%. In a phylogenetic tree all these isolates aphid species and estimate their numbers (Capote et al., 2008). All identified abundant aphid species collected from complete sticky rootstock plants were used to esti- mate the number of PPV-viruliferous aphids visiting the experimental nurseries in 2011. Aphids were squashed in- dividually on nylon membranes using the round bottom of an Eppendorf tube to ensure complete disruption of each aphid. RNA was extracted from the individual squashed aphid specimens using 100 μl buffer [0.1M glycine, 0.05 M NaCl, 1 mM ethylenediaminetetraaceticacid (EDTA)] (Osman and Rowhani, 2006) and analyzed by squash real- time RT-PCR (Olmos et al., 2005).

RESULTS

Evaluation of the susceptibility of different Prunus rootstocks to natural PPV-T infection. Individual root- stocks of the experimental plot established in 2009 were tested by DAS-ELISA in April and September of 2010 and 2011. Infection rate in 2010 was 6.0% and 4.0% for Nemaguard and Myrobalan 29C, respectively. It increased to 16.0% for Nemaguard and 14.0% for Myrobalan 29C Fig. 2. Phylogenetic analysis of PPV isolates from Myrobolan in 2011. The other four rootstocks were PPV-negative in 29C (TR-331, 332, 334, 336, 337, 338), Nemaguard (TR-329, these tests (Table 1). The most obvious symptoms on the 330, 333) in experimental plot and six apricot plants in a PPV- two infected rootstocks were vein clearing and interveinal infected orchard (TR-157ap, TR-159ap, TR-179ap, TR-192ap, TR-218ap, TR-220ap). Reference sequences were retrieved chlorosis (Fig. 1). Due to Turkish quarantine regulations, from GenBank, the first capital letters are accession numbers the PPV-infected orchard and all rootstocks were eradi- followed by isolate names. The percentage of replicate trees in cated three years after the establishment of the experi- which the associated taxa clustered together in the bootstrap mental plots. test (1000 replicates) is shown next to the branches.

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CP (Candresse et al., 2011). CP sequence alignments of PPV-T isolates with other reference isolates showed that this protein motif was also conserved in all sequenced iso- lates (Fig. 3).

Estimation of aphid species and number of virulifer- ous aphids present in the experimental plot. The aphid species landing on the rootstocks in the experimental plot were identified during spring (from April to June) of 2010 and 2011. A total of 73 and 364 individual aphids were captured in 2010 and 2011, respectively. M. persicae and H. pruni were the most common aphids in both years, fol- lowed by A. craccivora and A. gossypii (Table 2). In both years the total aphid number reached a peak at the end of May (26th of May in 2010, 31th of May in 2011). The most preferred rootstock by aphids was Nemaguard, followed by P. marianna GF 8.1, Garnem and Myrobalan 29C in both years (Fig. 4). Among all aphid species collected in 2011, the high- est percentage of viruliferous individuals was detected in Myzus persicae (34.78%) by squash real-time RT-PCR, followed by H. pruni (32.50%), Macrosiphum euphorbiae Fig. 3. Local alignment of Plum pox virus coat protein amino (25.00%), A. gossypii (23.80%), A. spiraecola (12.50%) and acid sequences. The isolates beginning with TR- are PPV-T A. craccivora (10.00%) (Table 3). An average of 24.19% isolates sequenced in this study, others were retrieved from (30 out of 124 analyzed individuals) of the aphid species GenBank. The conserved protein motif is shaded. that visited the experimental plot was viruliferous in the period studied. clustered in the PPV-T group (Fig. 2). As it can be seen The highest percentage of viruliferous aphids was in Fig. 2, the isolate AbTk was slightly different from the found on the rootstocks they preferred the most, i.e. My- other isolates obtained from the experimental nursery robalan 29C, P. marianna GF 8.1, Garnem and Nema- and the apricot orchard. Neverthless, the Mab 5B-IVIA guard (Fig. 3, Table 3). The two most common aphid spe- reacted with all PPV-T isolates. The reactivity of Mab 5B- cies, M. persicae and H. pruni, had the highest percentage IVIA with all PPV strains was investigated in a PPV ring of viruliferous individuals. test performed in the frame of a COST-88 concerted ac- tion (Asensio, 1996). Its positive reaction was also proved for strains PPV-EA, PPV-SoC and PPV-W (Wetzel et al., DISCUSSION 1991; Nemchinov and Hadidi, 1996; James and Varga, 2005). Mab 5B-IVIA reacts with the single protein mo- The PPV isolates found in the apricot orchard (inocu- tif (DRDVDAG sequence) fully conserved in PPV-D and lum source) and in the rootstocks of the experimental plot PPV-M, corresponding to positions 94 to 100 of the PPV were of the PPV-T type, which is a common strain in old

70 140 Garnem Garnem

60 Myrobolan 120 Myrobalan 29C GF 677 GF 677 50 Nemaguard 100 Nemaguard ds ds P.marianna GF 8.1 P.marianna GF 8.1 hi hi 40 80 ap ap Docera 6 Docera 6 of of er er 30 60 numb numb 20 40

10 20

0 0

l

l ta ta 21 , 28 , 14 , 07 , 02 , 11 , 26 , l 03 , 10 , 17 , 24 , 31 , 05 , 12 , 19 , 26 ,

l To l To ri y y y y y y y y y ri ri 2011 2011 2011 2011 2011 2011 2011 2010 2010 2010 2010 2010 2010 2010 2011 2010 une une une une Ap J J J J Ma Ma Ma Ma Ma Ap Ma Ma Ma Ma Ap

date date Fig. 4. Occurrence of PPV vector aphid species in different Prunus rootstocks. Number of alate individuals caught in spring of 2010 and 2011 (April–June) by the sticky plant method.

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stone fruit trees in Turkey. It is known that different PPV In this study, under severe PPV inoculum pressure the strains differ in terms of epidemiology and aggressiveness, assayed rootstocks exhibited differences in their suscepti- PPV-M is more severe and more efficiently vectored than bility to PPV-T infection by aphids (Table 1). Nemaguard PPV-D (Wang et al., 2006), thus it can spread very rap- and Myrobolan 29C proved highly susceptible, with in- idly in peach orchards (Dallot et al., 1998). PPV-D isolates fection rates of 16.0% and 14.0% in 2011, respectively, spread naturally in apricot and plum orchards but much in agreement with results from similar studies (Vidal et more slowly from these hosts to peach trees (Quiot et al., al., 2010). The susceptibility observed in Nemaguard, an 1995; Cambra et al., 2008). Limited information is available interspecific cross between peach and P. davidiana, agrees on the epidemic behavior and aggressiveness of PPV-Rec with the results reported by Pascal et al. (2002) and Ru- isolates and comparable data are lacking for PPV-T. The bio et al. (2003), indicating a certain level of susceptibility results of vector transmission studies (Glasa et al., 2004a) to a PPV-M and -D isolates, respectively. Although it is confirm that all the recombinant PPV isolates are aphid- known that P. cerasifera is susceptible to PPV (James and transmitted, but transmission occurrs at different rates. Thompson, 2006), different clones have been reported as

Table 2. Occurrence of PPV-vector aphid species in the experimental plot in Izmir. Number of individuals caught and their per- centage among total aphid numbers in the seasons of 2010 and 2011 (April-June).

2010 2011 Aphid species Number of Number of identified aphid Percentage (%) identified aphid Percentage (%) Myzus persicae 20 27.39 47 12.91 Hyalopterus pruni 17 23.28 51 14.01 Aphis craccivora 7 9.58 31 8.51 Aphis gossypii 6 8.21 31 8.51 Acyrthosiphon pisum 6 8.21 0 0 Anoecia corni 5 6.84 0 0 Aphis fabae 0 0 23 6.31 Macrosiphum euphorbiae 4 5.47 21 5.76 A. spiraecola 0 0 21 5.76 Hyperomyzus lactucae 4 5.47 0 0 Metopolophium dirhodum 3 4.10 0 0 Capitophorus elaeagni 1 1.36 0 0 helichrysi 0 0 4 1.09 Others 0 0 135 37.08 Total 73 364

Table 3. The number of viruliferous aphids/total number of aphids of that species collected from different Prunus rootstocks present in the experimental plot in 2011.

Garnem Myrobalan 29C Nemaguard GF677 P. marianna GF 8.1 Total 8/23 Myzus persicae 1/2 2/3 2/9 0/4 3/5 34.78% 13/40 Hyalopterus pruni 3/8 6/15 2/8 0/5 2/4 32.50% 1/4 Macrosiphum euphorbiae 0/0 0/0 1/4 0/0 0/0 25.00% 5/21 Aphis gossypii 2/6 2/4 1/7 0/3 0/1 23.80% 1/8 A. spiraecola 0/1 0/3 0/2 0/0 1/2 12.50% 2/20 A. craccivora 0/1 0/0 0/0 1/10 1/9 10.00% 0/8 A. fabae 0/2 0/0 0/1 0/4 0/1 0.00 6/20 10/25 6/31 1/26 7/22 30/124 Total 30.00% 40.00% 19.35% 3.84% 31.81% 24.19%

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resistant to PPV-D (Minoiu et al., 1998; Rubio et al., 2005). be done to determine their role in the epidemiology of In our experiment Myrobalan 29C was found as sensitive PPV-T in Turkey. as Nemaguard to PPV-T but the symptoms on this root- stock were always much milder than on Nemaguard (Fig. 1). Although some studies indicated the high susceptibility ACKNOWLEDGEMENTS of different clones of “Marianna” plum to PPV-M (Dosba et al., 1994), PPV-C (Bodin et al., 2003) and PPV-D (Rubio The research leading to these results received fund- et al., 2005), no infected Marianna 8.1 plants were detected ing from the European Community’s Seven Framework in this study, either because of low inoculum pressure or Programme (FP7/2007-2013) under Grant Agreement presence of different PPV strains. n°204429, SharCo project. The authors would like to PPV isolates from nursery and adjacent orchard that thank R. R. Martin from USDA-ARS Horticulture Crops were sequenced in this study were not 100% identical. Research Unit for his kind help for English editing. The rapidly evolving ability of plant RNA viruses has a significant role in virus epidemiology, as it provides a se- lection of variants with increased pathogenicity (Moury REFERENCES et al., 2006). Jiridi et al. (2006) demonstrated that 15 years after inoculation of a peach seedling with PPV-M, the vi- Ali A., Li H., Schneider W.L., Sherman D.J., Gray S., Smith D., rus could evolve into several distinct populations follow- Roossinck M.J., 2006. Analysis of genetic bottlenecks during horizontal transmission of Cucumber mosaic virus. Journal ing the systemic invasion of the host. In a plum tree triply of Virology 80: 8345-8350. infected with PPV-M, PPV-D and PPV-Rec, after seven Asensio M., 1996. El virus de la Sharka (plum pox virus): Car- years only the more competitive isolate PPV-M was still acterizacion, diagnóstico y detección mediante anticuerpos detectable whereas the two other isolates (PPV-Rec and monoclonales especificos. Ph.D. Thesis. Universidad Poli- PPV-D) had been displaced (Predejna et al., 2012). On the técnica de Valencia, Valencia, Spain. other hand, nonpersistent transmission by aphids may have Avinent L., Hermoso de Mendoza A., Llácer G., 1993. Com- an important effect on the dynamics and evolution of virus parison of sampling methods to evaluate aphid populations populations, for example a major cause of virus strain dif- (Homoptera, Aphidinea) alighting on apricot trees. Agrono- ferentiation could be genetic drift as a result of population mie 13: 609-613. bottlenecks during aphid transmission (Ali et al., 2006). Barba M., Hadidi A., Candresse T., Cambra M., 2011. Plum pox For epidemiological purposes, it is important to take virus. In: Hadidi A., Barba M., Candresse T., Jelkmann W. into consideration the aphid vector species as well as the (eds). Virus and Virus-like Diseases of Pome and Stone number of viruliferous aphids visiting the nursery plots. Fruits, pp. 185–197. APS Press, St. Paul, MN, USA. The most important PPV vectors reported from several Bodin, M., Glasa M., Verger D., Costes E., Dosba F., 2003. Dis- countries are Brachycaudus cardui, B. helichrysi, M. persi- tribution of the sour cherry isolate of Plum pox virus in in- cae and Phorodon humuli (Sullivan, 2011) and A. spirae- fected Prunus rootstocks. Journal of Phytopathology 151: 625- 630. cola (Cambra et al., 2006). Natural virus spread is low in Caglayan K., Kaya K., Serce Ç.U., Gazel M., Elci E., Cengiz summer but high in spring and autumn. Spring flights F.C., Vidal E., Cambra M., 2012. Epıdemıology of Plum pox of B. helichrysi, M. persicae, and P. humuli are the most vırus-T and -M isolates in stone fruit orchards in Turkey. important for virus spread within and between orchards Proceedings 22nd International Conference on Virus and Other (Sullivan, 2011). In our conditions, the total aphid popula- Graft Transmissible Diseases of Fruit Crops, Rome, Italy: 34. th tion reached a peak at the end of May in both years (26 Cambra M., Asensio M., Gorris M.T., Perez E., Camarasa E., of May in 2010, 31th of May in 2011). Garcia J.A., Moya J.J., Lopez-Abella D., Vela C., Sanz A., Tables 2 and 3 show that the most abundant aphid 1994. Detection plum pox potyvirus using monoclonal an- species were M. persicae and Hyalopterus pruni, and that tibodies to structural and non-structural proteins. OEPP these two species had the highest percentage of virulifer- Bulletin/EPPO Bulletin 24: 569-577. ous aphids. Although M. persicae is a well known PPV Cambra M., Capote N., Cambra M.A., Llacer G., Botella P., vector in many stone fruit-growing countries (Manachini Lopez A., 2006. Epidemiology of sharka disease in Spain. et al., 2007), the role of H. pruni in the natural spread of OEPP Bulletin/EPPO Bulletin 36: 271-275. PPV needs a better substantiated evidence (Gaborjanyi Cambra M., Flores R., Pallás V., Gentit P., Candresse T., 2008. and Basky, 1995). Of the aphids that visited the experi- Viruses and viroids of peach trees. In: Layne D.R., Bassi D. (eds). The Peach: Botany, Production and Uses, pp. 435-466. mental plot in Izmir, 24.19% (30 out of 124 analyzed in- CABI, Wallinford, UK. dividuals) were viruliferous in the period studied, which Candresse T., Cambra M., Dallot S., Lanneau M., Asensio M., explains the high incidence and rapid spread of PPV in Gorris M.T., Revers F., Macquaire G., Olmos A., Boscia D., the experimental plot in the studied area and the high risk Quiot J.B., Dunez J., 1998. Comparison of monoclonal anti- of PPV dissemination in stone fruit orchards. Most of the bodies and polymerase chain reaction assay for the typing of aphid species that were previously reported as PPV vectors isolates belonging to the D and M serotypes of plum pox were also able to transmit PPV-T but more work needs to potyvirus. Phytopathology 88: 198-204.

19. JPP1641RP (Caglayan).indd 584 06/11/13 16:10 Journal of Plant Pathology (2013), 95 (3), 579-586 Caglayan et al. 585

Candresse T., Cambra M., 2006. Plum pox virus strains or types: James D., Varga A., 2005. Nucleotide sequence analysis of Plum historical perspective and current status. OEPP Bulletin/ pox virus isolate W3174: evidence of a new strain. Virus Re- EPPO Bulletin 36: 239-246. search 110: 143-150. Candresse T., Svanella-Dumas L., Gentit P., Çaglayan K., Çevik Hall T.A., 1999. BioEdit: a user-friendly biological sequence B., 2007. First report of the presence of Plum pox virus Rec alignment editor and analysis program for Windows 95/98/ strain in Turkey. Plant Disease 91: 331. NT. Nucleic Acids Symposium Series 41: 95-98. Candresse T., Saenz P., Garcia J.A., Boscia D., Navratil M., Jridi C., Martin J-F., Marie-Jeanne V., Labonne G., Blanc S., Gorris M.T., Cambra M., 2011. Analysis of the epitope struc- 2006. Distinct viral populations differentiate and evolve in- ture of Plum pox virus coat protein. Phytopathology 101: 611- dependently in a single perennial host plant. Journal of Virol- 619. ogy 80: 2349-2357. Capote N., Perez-Panades J., Monzo C., Carbonell E.A., Ur- Kurcman S., 1973. Nachweis des sharka-virus an aprikosen und baneja A., Scorza R., Ravelonandro M., Cambra M., 2008. pflaumenbaumen aumenbaumen in Ankara. Journal of Turk- Assessment of the diversity and dynamics of Plum pox virus ish Phytopathology 2: 124-129. and aphid populations in transgenic European plums under Manachini B., Casati P., Cinanni L., Bianco P., 2007. Role of Mediterranean conditions. Transgenic Research, 17: 367-377. Myzus persicae (: ) and its secondary Dallot S., Labonne G., Quiot-douine L., Boeglin M., Candres- hosts in Plum pox virus propagation. Journal of Economic se T., Quiot J.B., 1998. Peculiar Plum pox potyvirus D pop- Entomology 100: 1047-1052. ulations are epidemic in peach trees. Acta Horticulture Marroquin C., Olmos A., Gorris M.T., Bertolini E., Martinez 472: 355-365. M.C., Carbonell E., Hermoso de Mendoza A., Cambra M., Deborré G., Maiss E., Jelkman W., 1995. Biological and mo- 2004. Estimation of the number of aphids carrying Citrus triste- lecular investigations of several Plum pox virus (PPV) iso- za virus that visit adult citrus trees. Virus Research 100: 101-108. lates. Acta Horticulturae 386: 253-262. Minoiu M., Maxim A., Vladianu D., Platon I., Balaci R., 1998. Dosba F., Lansac M., Eyquart J.P., Bonet A., Salesses G., 1994. New results concerning the Plum pox virus epidemiology Behaviour towards PPV of Prunus interspecific hybrids and and resistance of plum cultivars, hybrids and rootstocks. plum varieties. Acta Horticulturae 359: 136-144. Acta Virologica 42: 244-247. EPPO, 2004. Diagnostic protocol for regulated pests. Plum pox Moury B., Desbiez C., Jacquemond M., Lecoq H., 2006. Ge- potyvirus. OEPP Bulletin/EPPO Bulletin 34: 247-256. netic diversity of plant virus populations: towards hypoth- Gaborjanyi R., Basky S., 1995. Correlation between migration esis testing in molecular epidemiology. Advances in Virus of aphid vector and natural spread of plum pox virus. Acta Research 67: 49-87. Horticulturae 386: 201-206. Myrta A., Di Terlizzi B., Boscia D., Caglayan K., Gavriel I., Gazel M., Serce C.U., Caglayan K., 2010. New outbreaks of Ghanem G., Varveri C., Savino V., 1998. Detection and se- Plum pox virus in Turkey SharCo Research Workshop, Sofia, rotyping of Mediterranean Plum pox virus isolates by means Bulgaria: 27. of strain-specific monoclonal antibodies. Acta Virologica 42: 251-253. Glasa M., Boeglin M., Labonne G., 2004a. Aphid transmission of natural recombinant Plum pox virus isolates to different Nemchinov L., Hadidi A., 1996. Characterization of the sour Prunus ssp. - A contribution for understanding the epidemi- cherry strain of Plum pox virus. Phytopathology 86: 575-580 ology of an atypical PPV. Acta Horticulturae 657: 217-220. Olmos A., Bertolini E., Gil M., Cambra M., 2005. Real-time Glasa M., Palkovics L., Komínek P., Labonne G., Pittnerová S., assay for quantitative detection of non-persistently transmit- Kúdela O., Candresse T., Pubr Z., 2004b. Geographically ted Plum pox virus RNA targets in single aphids. Journal of and temporally distant natural recombinant isolates of Plum Virological Methods 128: 151-155. pox virus are genetically very similar and form a unique PPV Osman F., Rowhani A., 2006. Application of a spotting sample subgroup. Journal of General Virology 85: 2671-2681. preparation technique for the detection of pathogens in Glasa M., Candresse T., 2005. Partial sequence analysis of an woody plants by RT-PCR and real-time PCR (TaqMan). Jour- atypical Turkish isolate provides further information on the nal of Virological Methods 133: 130-136. evolutionary history of Plum pox virus (PPV). Virus Research Palmisano F., Bazzoni A., Tavano D., Didonna A., Castellano 108: 199-206. M., Savino V., 2010. Experimental screening of an apricot Gümüs M., Paylan I.C., Matic S., Myrta A., Sipahioglu H.M., cross for resistance to Plum pox virus (M and D strains). Erkan S., 2007. Occurrence and distribution of stone fruit Acta Horticulturae 862: 501-506. viruses and viroids in commercial plantings of Prunus spe- Pascal T., Pfeiffer F., Kervella J., 2002. Preliminary observations cies in western Anatolia, Turkey. Journal of Plant Pathology on the resistance to Sharka in peach and related species. 89: 265-268. Acta Horticulturae 592: 699-704. James D., Thompson D., 2006. Hosts and symptoms of Plum Predajna L., Subr Z., Candresse T., Glasa N., 2012. Evaluation pox virus: ornamental and wild Prunus species. Bulletin of the genetic diversity of Plum pox virus in a single plum OEPP/EPPO Bulletin 36: 222-224. tree. Virus research 167: 112-117 Jarausch W., Baßler A., Molla N., Krczal G., 2004. First detec- Quiot J.B., Boeglin M., Admolle C., Candresse T., Labonne G., tion and molecular characterisation of PPV-M strains in Renaud L.Y., 1995. Behaviour of two isolates of Plum pox plum orchards in South-Western Germany. Acta Horticul- virus inoculated on peach and apricot trees: First results. turae 657: 159-164. Acta Horticulturae 386: 290-297.

19. JPP1641RP (Caglayan).indd 585 06/11/13 16:10 586 Susceptibility to infection of Plum pox virus-T Journal of Plant Pathology (2013), 95 (3), 579-586

Rubio M., Martinez-Gomez P., Dicenta F., 2003. Resistance of USDA-APHIS-PPQCPHST. almond cultivars to Plum pox virus (Sharka). Plant Breeding Tamura K., Peterson D., Peterson N., Stecher G., Nei M., Ku- 122: 462-464. mar S., 2011. MEGA5: Molecular evolutionary genetics Rubio M., Martınez-Gomez P., Pinochet J., Dicenta F., 2005. analysis using maximum likelihood, evolutionary distance, Evaluation of resistance to sharka (Plum pox virus) of sev- and maximum parsimony methods. Molecular Biology and eral Prunus rootstocks. Plant Breeding 124: 67-70. Evolution 28: 2731-2739. Ulubas Serce C., Candresse T., Svanella-Dumas L., Krizbai L., Vidal E., Moreno A., Bertolini E., Perez-Panades J., Carbonell Gazel M., Caglayan K., 2009. Further characterization of a E.A., Cambra M., 2010. Susceptibility of Prunus rootstocks new recombinant group of Plum pox virus isolates, PPV-T, to natural infection of Plum pox virus and effect of mineral found in orchards in the Ankara province of Turkey. Virus oil treatments. Annals of Applied Biology 157: 447-457. Research 142: 121-126. Wang A., Sanfacon H., Stobbs L.W., James D., Thompson D., Sahtiyancı S., 1969. Virus de la sharka chez le prunier. Bulletin Svircev A.M., Brown D.C.W., 2006. Plum pox virus in Can- Phytosanitaire FAO 17: 69. ada: progress in research and future prospects for disease Spiegel S., Scott S.W., Bowman-Vance V., Tam Y., Galiakparov control. Canadian Journal of Plant Pathology 28: 182-196. N.N., Rosner A., 1996. Improved detection of Prunus ne- Wetzel T., Candresse T., Ravelonandro M., Delbos R.P., Mazyad crotic ringspot virus by polymerase chain reaction. European H., Aboul-Ata E.E., Dunez J., 1991. Nucleotide sequence of Journal of Plant Pathology 102: 681-685. the 3’-terminal region of the RNA of the El Amar strain of Sullivan M., 2011. CPHST Pest Datasheet for Plum pox virus. plum pox potyvirus. Journal of General Virology 72: 1741-1746.

Received March 15, 2013 Accepted May 16, 2013

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