First Report of an Elm Yellows Subgroup 16SrV-C Phytoplasma Infecting Grapevine in Serbia

During a 2002 survey in Serbia, samples of grapevine (Vitis vinifera) were collected from plants showing typical phytoplasma-like symptoms: leaf roll, leaf redness, vein chlorosis and necrosis, and absence of lignification. The material was collected from one viticultural region (Zupa Aleksandrovac), where the disease was recorded in 2000 and showed an increasing percentage of symptomatic plants every year. Total nucleic acid was extracted separately from leaf midveins and stem bark collected from 10 symptomatic and 2 asymptomatic plants. Phytoplasma infection was detected using polymerase chain reaction (PCR) assays with universal primer pair P1/P7 for the amplification of phytoplasma 16S rRNA gene, and primer pair FD9f2/FD9r followed by FD9f3/FD9r2 in nested PCR for specific amplification of the FD9 nonribosomal DNA fragment of the EY-group (1). Phytoplasmas were detected in 9 of 10 midvein extracts from symptomatic grapevines (three of cv. Plovdina, two of cv. Smederevka, and four of cv. Gamé). Also, 6 of 10 bark preparations representing stem collections from the same plants were positive (two samples of cv. Plovdina, both samples of cv. Smederevka, and two samples of cv. Gamé). Both collections of midveins and bark tissues from asymptomatic plants were negative. Fragments amplified with universal P1/P7 primers (16S-23S rDNA) were analyzed by restriction fragment length polymorphism with TruI and TaqI restriction enzymes. The phytoplasmas produced identical restriction profiles to those of 16SrV Elm Yellows group and 16SrV-C Flavescence doreé subgroup (2). To our knowledge, this is the first report of phytoplasma infecting grapevines in Serbia, and the first survey in progress to verify the presence of Scaphoideus titanus to determine if this grapevine yellows could be defined as Flavescence dorée. References: (1) E. Angelini et al. Vitis 40:79, 2001. (2) M. Martini et al. Mol. Cell. Probes 16:197, 2002.

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First report of Flavescence Dorée-Related Phytoplasma in a Productive Vineyard in Germany

Plant Disease ◽

10.1094/pdis-02-21-0330-pdn ◽

2021 ◽

Author(s):

Barbara Jarausch ◽

Sandra Biancu ◽

Sanela Kugler ◽

Thierry Wetzel ◽

Manuel Baumann ◽

...

Keyword(s):

Real Time ◽

Single Case ◽

Rrna Gene ◽

The European Union ◽

First Report ◽

Flavescence Dorée ◽

Black Alder ◽

Grapevine Yellows ◽

Scaphoideus Titanus ◽

Infected Vine

Flavescence dorée (FD) and Bois noir (BN) are the principal grapevine yellows in Europe caused by distinct phytoplasmas: BN by Candidatus Phytoplasma solani, FD by 16SrV-C and -D phytoplasmas (FDp) transmitted by the introduced Nearctic Deltocephalinae Scaphoideus titanus. FDp is listed as a quarantine pest in the European Union (Regulation (EU) 2019/2072). Black Alder (Alnus glutinosa) is a common asymptomatic host of 16SrV phytoplasmas in Europe and considered the original host of FDp (Malembic-Maher et al. 2020). Palatinate grapevine yellows (PGY) transmitted from alder to grapevine by the Macropsinae Oncopsis alni is not transmissible by S. titanus (Malembic-Maher et al. 2020). Germany is considered free from FD in grapevine and from its vector. A single case in a nursery in 2014 was eradicated (EPPO 2017), and FD was never before detected in a vineyard. Since S. titanus appeared in 2016 in the neighboring French Region of Alsace, monitoring of FD was carried out in Germany following a risk based strategy. It was focused on vineyard plots within a distance of 100 m from stands of alder. A geodata-based risk map (Jalke 2020) was used to localize those plots. All symptomatic vines sampled until September 2020 proved to be infected by BN or, occasionally, by PGY. Eight vines with typical symptoms were sampled in vineyards adjacent to alder stands in the winegrowing region of Rheinhessen in September 2020. Symptoms comprised leaf rolling and discoloration, incomplete lignification, and black pustules arranged in lines along the shoots. Diseased shoots were black and necrotic in December. Leaf midribs were sampled for total nucleic acids extraction. The phytoplasma 16S rRNA gene was amplified by generic primers R16F2/R2-mod followed by a nested PCR using 16Sr(V) group-specific primers R16(V)F1/R1, and primers R16(I)F1/R1 (Lee et al. 1995) to detect ‘Candidatus Phytoplasma solani’, associated with BN. While BN was detected in seven vines, one sample tested positive for 16SrV phytoplasma. This result was confirmed by triplex real-time Taq-Man assay based on rpl14 gene sequences (IPADLAB), by multiplex real-time PCR of map locus as well as by Loop-mediated isothermal amplification (LAMP) according to the EPPO diagnostic standard PM 7/079(2) (EPPO 2016). PCR-products of the map and the vmpA genes (Malembic-Maher et al. 2020) were sequenced and compared to reference sequences to distinguish between FD- and non-FD genotypes. The isolate from the diseased vine (GenBank MW 727272) exhibited 100% identity with map-M38 (GenBank LT221933), a genotype of the map-FD2 cluster. The same genotype was detected in A. glutinosa and Allygus spp. sampled at the infested site. A 234 bp sequence of the first repeat of the vmpA gene (GenBank MW727273) showed 100% identity with the homologous part of isolate FD-92 (GenBank LN680870) of the vmpA-II cluster. It can be concluded, that the symptomatic grapevine was infected by FD and not PGY This is the first report of FD in a productive vineyard in Germany. The infected vine of cv. Silvaner was 25 years old. While infected planting material is an unlikely source of the infection, a transmission of FDp from alder is highly probable. Finding a single FD-infection after several years of testing implies a low risk originating from the wild compartment, but the approach and possible establishment of S. titanus expected to be able to colonize the area (Jeger et al. 2016) justifies further monitoring activities. The infected vine was eradicated.

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First Report of Alder Yellows Phytoplasma Associated with Common Alder (Alnus glutinosa) in the Republic of Macedonia

Plant Disease ◽

10.1094/pdis-03-14-0315-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1268-1268 ◽

Cited By ~ 5

Author(s):

B. Atanasova ◽

D. Spasov ◽

M. Jakovljević ◽

J. Jović ◽

O. Krstić ◽

...

Keyword(s):

Genetic Locus ◽

Ribosomal Protein Gene ◽

Alnus Glutinosa ◽

Sensu Stricto ◽

Universal Primers ◽

Rrna Gene ◽

Significant Similarity ◽

First Report ◽

Flavescence Dorée ◽

Scaphoideus Titanus

Alder yellows phytoplasma (AldYp) is classified as a member of the 16SrV-group of phytoplasmas and is closely related to Flavescence dorée (FD), a quarantined pathogen of economic importance affecting vineyards across Europe. AldYp is associated with common (Alnus glutinosa) and grey alder (A. incana), and has been reported in France, Italy, Germany, Austria, Switzerland, the Baltic region, Serbia, and Montenegro (1,2,4). For Macedonian vineyards, so far, neither infection of grapevine with 16SrV-group of phytoplasmas nor the presence of the main FD phytoplasma vector, Scaphoideus titanus, has been recorded. However, the presence of FD-related phytoplasma was detected in wild Clematis vitalba. In September and October 2013, leaves with petioles from A. glutinosa exhibiting leaf discoloration and yellowing were collected from two sites (41°23′43″ N, 22°54′ E and 41°23′ N, 22°53′ E) in southeast Macedonia near the village of Smolare (Strumica district). Eight samples were collected from each site. Leaves of six asymptomatic alder seedlings collected from the same sites served as a control. Nucleic acids were extracted from fresh leaf midribs and petioles using a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany). Initial phytoplasma identification was carried out by nested PCR assay of the 16S rRNA gene, using universal primers P1/P7 and R16F2n/R16R2 followed by RFLP with MseI endonuclease (Fermentas, Vilnius, Lithuania), as previously reported (4). Characterization of detected phytoplasmas was performed by amplifying two genetic loci specific for the members of the 16SrV group phytoplasmas; the ribosomal protein gene operon (rp) using primers rp(V)F1/rpR1 and rp(V)F1A/rp(V)R1A (3), and the non-ribosomal metionine aminopeptidase (map) gene using primer set FD9f5/MAPr1 and FD9f6/MAPr2 (1). The PCR amplicons were sequenced and deposited in NCBI GenBank database under the accession numbers KJ605448 to 52 (map) and KJ605453 to 57 (rp). The obtained sequences were compared with reference sequences of the 16SrV-group phytoplasmas (1,3) using the neighbor-joining method in MEGA5 (5). The presence of phytoplasma was detected in 14 of 16 symptomatic alder samples, while all control plants tested negative. The MseI restriction profiles were identical among all 14 samples and with the reference strains of the 16SrV group phytoplasmas (EY1 - 16SrV-A, FD-C - 16SrV-C, and FD-D - 16SrV-D). The rp-based phylogeny enabled identification of four diverse phytoplasma strains among the AldYp strains from Macedonia. Three strains clustered within the rpV-E subgroup while one belonged to rpV-L subgroup. Phylogenetic analysis of the more variable genetic locus, map, showed the presence of five diverse phytoplasma strains. Four strains belonged to the map-FD2 (FD-D, FD92) cluster, while one grouped within the map-FD1 (FD70) cluster. To our knowledge, this is the first report of 16SrV phytoplasma group occurrence on alder in Macedonia. The significant similarity between AldYp strains and FD sensu stricto indicate the risk of pathogen exchange between the wild ecosystem and the grapevine (1). Alder trees naturally infected with the FDp-related strains could therefore represent a serious risk for FD outbreak in Macedonian vineyards if local S. titanus populations developed. References: (1) G. Arnaud et al. Appl. Environ. Microbiol. 73:4001, 2007. (2) T. Cvrković et al. Plant Pathol. 57:773, 2008. (3) M. Martini et al. Int. J. Syst. Evol. Microbiol. 57:2037, 2007. (4) S. Radonjić et al. Plant Dis. 97:686, 2013. (5) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011.

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First Report of Elm Yellows Subgroup 16SrV-B Phytoplasma Infecting Chinese Tulip Tree in China

Plant Disease ◽

10.1094/pdis-03-12-0281-pdn ◽

2012 ◽

Vol 96 (7) ◽

pp. 1064-1064 ◽

Cited By ~ 2

Author(s):

Z. Y. Li ◽

Z. P. Dong ◽

Z. M. Hao ◽

J. G. Dong

Keyword(s):

Sequence Data ◽

Leaf Roll ◽

Universal Primer ◽

Group V ◽

First Report ◽

Blast Analysis ◽

Slow Decline ◽

Specific Fragment ◽

Elm Yellows ◽

Tulip Tree

Chinese tulip tree (Liriodendron chinensis) is native to China and is planted all around the country as an ornamental tree. In July of 2011, some Chinese tulip trees with typical phytoplasma symptoms were found in Baoding City, Hebei Province, China. Symptoms included yellowing of leaves, slow decline, little leaves, and death of entire plants. To confirm phytoplasma infection of these plants, total DNA was extracted from 100 mg of fresh leaf midribs collected from leaves of nine symptomatic and eight asymptomatic plants with a plant DNA extract kit (Tiangen, Beijing, China) according to the manufacturer's protocols. Using 16S rRNA phytoplasma universal primer pairs P1/P7 followed by R16F2n/R16R2, a nest PCR was carried out (1,2). The results showed that the phytoplasma was only detected in symptomatic samples by nested PCR, while the asymptomatic were negative. An approximate 1.2-kb specific fragment was obtained from the DNA of nine symptomatic plants, but no product was amplified from the leaves of eight healthy ones. The amplified products were cloned and sequenced. The sequence was deposited in GenBank Data Libraries under Accession No. JQ585925 and shared the highest homology of 99% with Puna chicory flat stem phytoplasma (GenBank Accession No. JN582266), Apricot leaf roll phytoplasma (GenBank Accession No. FJ572660), Jujube witches'-broom phytoplasma (GenBank Accession No. AY197661), and other elm yellows group phytoplasmas by BLAST analysis with that of other phytoplasmas from GenBank. Meanwhile, the sequence data was analyzed by iPhyClassifier software and the result showed that the 16S rDNA F2nR2 fragment was identical (similarity coefficient 1.00) to the reference patterns of 16Sr group V, subgroup B (GenBank Accession No.AB052876) (3). Combining the BLAST analysis in GenBank and the analysis of iPhyClassifier, we classified the phytoplasma causing Chinese tulip tree yellow leaves disease into subgroup 16SrV-B. To our knowledge, this is the first report of the 16SrVB group phytoplasmas infecting Chinese tulip tree in China. References: (1) I. M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) I. M. Lee et al. Int. J. Syst. Evol. Microbiol. 54:337, 2004. (3) Y. Zhao et al. Int. J. Syst. Evol. Microbiol. 59:2582, 2009.

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First Report of Flavescence Dorée-Related Phytoplasma Affecting Grapevines in Croatia

Plant Disease ◽

10.1094/pdis-09-10-0664 ◽

2011 ◽

Vol 95 (3) ◽

pp. 353-353 ◽

Cited By ~ 7

Author(s):

M. Šeruga Musić ◽

D. Škorić ◽

I. Haluška ◽

I. Križanac ◽

J. Plavec ◽

...

Keyword(s):

Nested Pcr ◽

Reference Strain ◽

Simultaneous Detection ◽

Pinot Noir ◽

Vitis Vinifera L ◽

Rrna Gene ◽

Pcr Assay ◽

Direct Pcr ◽

Flavescence Dorée ◽

Scaphoideus Titanus

Flavescence dorée (FD) and Bois noir (BN) phytoplasmas are principal grapevine yellows (GY) agents in the wider Euro-Mediterranean Region. While BN phytoplasma belongs to the ribosomal subgroup 16SrXII-A, the FD agents belong either to the ribosomal subgroups 16SrV-C or -D. During the official GY survey in 2009, 40 symptomatic grapevines (Vitis vinifera L.) were sampled throughout grapevine-growing regions in Croatia. Typical GY symptoms of leaf yellowing or reddening were evident on white and red varieties, respectively. Leaf rolling as well as irregular lignification of the shoots and withering of clusters were also observed. Phloem tissue from cuttings and leaf veins from mature vines were sampled for total DNA extraction and amplification of phytoplasma 16S rRNA gene by using generic primers P1/P7 in a direct PCR assay followed by a nested PCR using primer pair R16F2n/R2 (2). Phytoplasma ribosomal group affiliation was determined by restriction fragment length polymorphism (RFLP) analysis of the nested PCR products with enzyme Tru1I (Fermentas, Vilnius, Lithuania). These initial findings were validated and augmented by a triplex real-time PCR assay targeting the nonribosomal map gene. This assay enables simultaneous detection of BN and FD (16SrV-C and -D) phytoplasmas in grapevine (3). Assay results revealed the majority of GY positive vines (19 of 40) contained BN phytoplasma which is widespread. For the first time in Croatia, two red variety samples, Pinot Noir and Plemenka Crvena, from the vicinity of Ozalj (Vivodina) and Zagreb (Brezje), respectively, were found to harbor FD-related phytoplasmas. Fragments amplified by P1/P7 primers from latter samples were cloned and sequenced. Sequence analyses using online interactive tool iPhyClassifier (4) revealed that the phytoplasma under study from Pinot Noir sample (GenBank Accession No. HQ712064) is a member of 16SrV-C subgroup and shares 99.87% similarity with 16S rDNA sequence of the reference strain (GenBank Accession No. AF176319). The sequence from the Plemenka Crvena sample (GenBank Accession No. HQ712065) shares 99.54% similarity with the reference strain and has the most similar virtual RFLP pattern to the one of the 16SrV-C subgroup (GenBank Accession No. AY197642). These findings are currently limited to vineyards in northwestern Croatia. Even so, the presence of FD principal cicadellid vector Scaphoideus titanus in the country and the occurrence and distribution of FD in neighboring countries (1,2) are factors indicating that the spread of FD in Croatia is highly probable. References: (1) L. Filippin et al. Plant Pathol. 58:826, 2009. (2) S. Kuzmanović et al. Vitis 47:105, 2008. (3) C. Pelletier et al. Vitis 48:87, 2009. (4) Y. Zhao et al. Int. J. Syst. Evol. Microbiol. 59:2582, 2009.

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Identification and Epidemic Distribution of Two Flavescence Dorée—Related Phytoplasmas in Veneto (Italy)

Plant Disease ◽

10.1094/pdis.1999.83.10.925 ◽

1999 ◽

Vol 83 (10) ◽

pp. 925-930 ◽

Cited By ~ 56

Author(s):

Marta Martini ◽

Ermanno Murari ◽

Nicola Mori ◽

Assunta Bertaccini

Keyword(s):

16S Rdna ◽

Fragment Length Polymorphism ◽

The European Union ◽

Direct Pcr ◽

Nested Polymerase Chain Reaction ◽

Flavescence Dorée ◽

Scaphoideus Titanus ◽

Polymerase Chain ◽

Elm Yellows ◽

Infected Grapevine

Grapevine yellows associated with phytoplasmas of the elm yellows group (16SrV), better known as flavescence dorée (FD), is a serious quarantine problem in some important grapevine growing regions in the European Union. A survey was carried out in 1997 to 1998 in Veneto region (Italy) where a serious outbreak of FD was in progress. Phytoplasma identification by nested polymerase chain reaction (PCR)/restriction fragment length polymorphism (RFLP) analyses on 275 grapevine samples and on batches of Scaphoideus titanus was carried out. RFLP analyses of the 16S rDNA/spacer region with TaqI detected the presence of two distinct elm yellows phytoplasma subgroups designated 16SrV-C and 16SrV-D in 77 FD-infected grapevine samples. Only phytoplasmas of the 16SrV-D subgroup were detected in S. titanus. In 1997, the two phytoplasma subgroups appeared to be located in two diverse geographic areas; but in 1998, the 16SrV-D type also was detected in the provinces where in 1997 only 16SrV-C type was identified. The sequencing of a 400-bp fragment at the 3′ end of 16S rDNA plus spacer region allowed a specific primer construction that was successfully employed for detection of both FD types in grapevine by direct PCR.

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First Report of Clover Proliferation Phytoplasma in Strawberry

Plant Disease ◽

10.1094/pdis.1999.83.10.967c ◽

1999 ◽

Vol 83 (10) ◽

pp. 967-967 ◽

Cited By ~ 4

Author(s):

R. Jomantiene ◽

J. L. Maas ◽

E. L. Dally ◽

R. E. Davis ◽

J. D. Postman

Keyword(s):

Dna Sequences ◽

Rrna Gene ◽

Universal Primer ◽

Disease Symptoms ◽

First Report ◽

Chain Reactions ◽

Polymerase Chain Reactions ◽

Fragaria Virginiana ◽

Phytoplasma Infection ◽

Group 2

In 1996, diseased plants of Fragaria virginiana Duchesne were collected from a native population in Quebec, Canada, and sent to the National Clonal Germplasm Repository in Corvallis, OR, where grafting onto disease-free plants of F. chiloensis (L.) Duchesne (4) was performed. Plants of both species were sent to Beltsville, MD, for identification of a phytoplasma possibly associated with the disease symptoms of dwarfing and multibranching crowns. A phytoplasma was found in both species and characterized as the strawberry “multicipita” (SM) phytoplasma, which is representative of subgroup 16SrVI-B, a new subgroup of the clover proliferation (CP) group (2). In 1999, we observed commercial strawberry (Fragaria × ananassa Duchesne) plants collected in California and Maryland that were stunted and chlorotic or exhibited these symptoms in addition to small, distorted leaves. Infected F. × ananassa plants, as well as diseased F. virginiana and grafted F. chiloensis plants previously infected by the SM phytoplasma, were assessed for phytoplasma infection by nested polymerase chain reactions primed by phytoplasma universal primer pairs R16mF2/R1 and F2n/R2 (1) or P1/P7 (3) and F2n/R2 for amplification of phytoplasma 16S rDNA (16S rRNA gene) sequences. Phytoplasma-characteristic 1.2-kbp DNA sequences were amplified from all diseased plants. No DNA sequences were amplified from healthy plants. Restriction fragment length polymorphism patterns of rDNA digested with AluI, KpnI, HhaI, HaeIII, HinfI, HpaII, MseI, RsaI, and Sau3A1 endonucleases indicated that all plants were infected by a phytoplasma that belonged to subgroup 16SrVI-A (CP phytoplasma subgroup) and that diseased F. virginiana and grafted F. chiloensis plants were infected by both SM and CP. This is the first report of the CP phytoplasma, subgroup 16SrVI-A, infecting strawberry. This report also indicates that the occurrence of the CP phytoplasma in strawberry may be widespread in North America and that F. chiloensis, F. virginiana, and F. × ananassa plants are susceptible to infection by the CP phytoplasma. References: (1) D. E. Gunderson and I.-M. Lee. Phytopathol. Mediterr. 35:144, 1996. (2) R. Jomantiene et al. HortScience 33:1069, 1998. (3) R. Jomantiene et al. Int. J. Syst. Bacteriol. 48:269, 1998. (4) J. D. Postman et al. Acta Hortic. 471:25, 1998.

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First Report of Pear Decline Phytoplasmas on Pear in Serbia

Plant Disease ◽

10.1094/pd-89-0774c ◽

2005 ◽

Vol 89 (7) ◽

pp. 774-774 ◽

Cited By ~ 3

Author(s):

B. Duduk ◽

M. Ivanović ◽

A. Obradović ◽

S. Paltrinieri ◽

A. Bertaccini

Keyword(s):

Nested Pcr ◽

Restriction Enzymes ◽

Diagnostic Procedures ◽

Nested Polymerase Chain Reaction ◽

Universal Primer ◽

First Report ◽

Taxonomy Group ◽

Pyrus Communis L ◽

Polymerase Chain ◽

Symptomatic Plant

During August of 2004, pear (Pyrus communis L.) plants with typical symptoms of pear decline (PD) were observed in orchards in central Serbia. The affected plants showed premature reddening and upward rolling of leaves that often showed down-turned petioles. In some cases, premature defoliation was observed. Although a similar decline of pear was observed earlier, until now, the causal agent had not been identified. DNA was extracted with a chloroform/phenol procedure from fresh leaf midribs and branch phloem scrapes of four symptomatic and one asymptomatic pear plants separately. A nested polymerase chain reaction assay (PCR) was used for phytoplasma detection (first PCR round with P1/P7 (4) phytoplasma universal primer pair, followed by nested PCR with group 16SrX specific primers f01/r01) (3). With these primers, the expected products from phloem scrapes and midrib extracts of symptomatic plant samples were obtained. Restriction fragment length polymorphism (RFLP) analyses of the f01/r01 amplicon, with RsaI and SspI restriction enzymes, discriminating among 16SrX subgroup phytoplasmas, showed profiles corresponding to those of the apple proliferation phytoplasma group, 16SrX-C subgroup, “Candidatus Phytoplasma pyri” (2). A 1,155-bp sequence of 16S rDNA gene for one of the PA2f/r (1) amplicons obtained in nested PCR on P1/P7 products from one of the leaf midrib samples was deposited in GenBank (Accession No. AY949984); both strands of the fragment were sequenced with the Big Dye Terminator reaction kit (Applied Biosystems, Foster City, CA). The sequences were analyzed with the Chromas 1.55 DNA sequencing software (Technelysium, Queensland, Australia) and aligned with BLAST software ( http://www.ncbi.nlm.nih.gov ). The blast search showed 100% homology of this sequence with that of PD strain Y16392, confirming the identity with PD of the phytoplasma detected. To our knowledge, this is the first report of pear decline phytoplasmas in Serbia. References: (1) M. Heinrich et al. Plant Mol. Biol. Rep. 19:169, 2001. (2) IRPCM Phytoplasma/Spiroplasma Working Team-Phytoplasma Taxonomy Group. Int. J. Syst. Evol. Microbiol. 54:1243, 2004. (3) K.-H. Lorenz et al. Phytopathology 85:771, 1995. (4) Schneider et al. Pages 369–380 in: Molecular and Diagnostic Procedures in Mycoplasmology. Vol I. S. Razin and J. G. Tully, eds. The American Phytopathological Society, 1995.

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Molecular Diversity of Phytoplasmas Associated with Grapevine Yellows Disease in North-Eastern Italy

Phytopathology ◽

10.1094/phyto-07-17-0253-r ◽

2018 ◽

Vol 108 (2) ◽

pp. 206-214 ◽

Cited By ~ 15

Author(s):

Yuri Zambon ◽

Alessandro Canel ◽

Assunta Bertaccini ◽

Nicoletta Contaldo

Keyword(s):

Molecular Diversity ◽

Direct Sequencing ◽

Northern Italy ◽

Bois Noir ◽

Aster Yellows ◽

Flavescence Dorée ◽

Scaphoideus Titanus ◽

North Eastern ◽

Elm Yellows ◽

Hyalesthes Obsoletus

A 3-year survey was conducted in Northern Italy to verify the presence and diversity of phytoplasmas in selected vineyards showing symptoms of severe yellows. Symptomatic and asymptomatic grapevines were sampled, and insects were collected using yellow sticky traps. The phytoplasmas detected in grapevine samples were different according to the years: “flavescence dorée” (16SrV-C/D) was detected together with other phytoplasmas such as 16SrXII-A (‘Candidatus Phytoplasma solani’-related, bois noir), 16SrI-B (‘Ca. P. asteris’-related, aster yellows), 16SrX-B (‘Ca. P. prunorum’-related, European stone fruit yellows), and 16SrV-A (‘Ca. P. ulmi’-related, elm yellows). Moreover, phytoplasmas belonging to 16SrVII-A (‘Ca. P. fraxini’-related) and 16SrVI (‘Ca. P. trifolii’-related) subgroups were also identified. Identification of phytoplasmas was also carried out from insects and showed the presence of some of these phytoplasmas in Scaphoideus titanus and Orientus ishidae: 16SrXII-A, 16SrVII, and 16SrVI phytoplasmas were detected in specimens of both species, while 16SrXII-A and 16SrI-B phytoplasma strains were identified in Orientus ishidae and Hyalesthes obsoletus, and 16SrX-B in S. titanus. Direct sequencing of selected amplicons obtained from 16S rRNA, rp, and tuf genes from grapevine and insect samples confirmed the phytoplasma identification. The 16SrVII-A and 16SrVI phytoplasmas were never detected before in grapevine, S. titanus and Orientus ishidae in Europe and their epidemiological importance is being monitored.

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Characterization of Phytoplasmas Related to Aster Yellows Group Infecting Annual Plants in Iran, Based on the Studies of 16S rRNA and RP Genes

Journal of Plant Protection Research ◽

10.2478/jppr-2014-0001 ◽

2014 ◽

Vol 54 (1) ◽

pp. 1-8 ◽

Cited By ~ 13

Author(s):

Fereshteh Vali Sichani ◽

Masoud Bahar ◽

Leila Zirak

Keyword(s):

16S Rrna ◽

Rflp Analysis ◽

Restriction Enzymes ◽

Rrna Gene ◽

Annual Plants ◽

Universal Primer ◽

Aster Yellows ◽

Central Regions ◽

Group Specific Primer

Abstract Several annual field crops, vegetables, ornamentals, oilseed crops, and weeds showing phytoplasma diseases symptoms were collected to detect phytoplasmas related to ‘Candidatus Phytoplasma asteris’. The collecting was done in the central regions of Iran. For general detection of phytoplasmas, 16S rRNA gene fragments were amplified using phytoplasma universal primer pair P1/P7 in polymerase chain reaction (PCR) followed by primer pair R16F2n/R16R2 in nested PCR. Then, for finer detection of phytoplasmas related to ‘Ca. P. asteris’, DNA samples were used to extend the rp and tuf gene fragments by PCR using aster yellows group specific primer pairs rp(I)F1A/rp(I)R1A and fTufAy/rTufAy, respectively. Restriction fragment lenght polymorphism (RFLP) analysis of rp gene fragments using digestion with AluI, MseI, and Tsp509I restriction enzymes indicated that aster yellows group related phytoplasmas in these Iranian regions, belong to rpI-B subgroups. Sequence analysis of partial 16S rRNA and rp genes from representative phytoplasma isolates confirmed the RFLP results. This research is the first report of annual plants infected with phytoplasmas related to subgroup rpI-B in Iran.

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First Report of Witches'-Broom Disease in a Cannabis spp. in China and Its Association with a Phytoplasma of Elm Yellows Group (16SrV)

Plant Disease ◽

10.1094/pdis-91-2-0227c ◽

2007 ◽

Vol 91 (2) ◽

pp. 227-227 ◽

Cited By ~ 11

Author(s):

Y. Zhao ◽

Q. Sun ◽

R. E. Davis ◽

I.-M. Lee ◽

Q. Liu

Keyword(s):

Nucleotide Sequence ◽

16S Rdna ◽

Geographic Location ◽

Full Length ◽

23S Rrna ◽

Nucleotide Sequence Analysis ◽

Rrna Gene ◽

Hemp Fiber ◽

First Report ◽

Elm Yellows

Hemp fiber plants (Cannabis spp.) spread naturally in almost every climate zone in China and have a long history of cultivation in the country (1). While hemp stalks provide high-quality fibers for making ropes, clothes, and paper products, hemp seeds are a rich source of edible oil. During the summer of 2004, a disease characterized by witches'-broom symptoms was observed in wild hemp fiber plants growing in suburban Taian, Shandong, China. The diseased plants developed clusters of highly proliferating branches with much shortened internodes and leaves on the affected branches were significantly reduced in size. Phytoplasma infection was suspected in this hemp fiber witches'-broom (HFWB) disease because of the typical symptoms and because of its geographic location where other phytoplasmal diseases such as jujube witches'-broom (JWB), paulownia witches'-broom (PaWB), paper mulberry witches'-broom (PMWB), and Chinese wingnut witches'-broom (CWWB) diseases were previously reported (3,4). Total DNA was extracted from leaves of four diseased and four nearby healthy looking hemp fiber plants. Nested PCR were carried out on the DNA samples using phytoplasma universal 16S rDNA primers (P1A/16S-SR and R16F2n/R16R2) (2). Results revealed that all examined diseased plants were infected by phytoplasma, whereas nearby healthy looking plants were phytoplasma free. Subsequent restriction fragment length polymorphism (RFLP) analysis of the PCR-amplified 1.25-kb 16S rDNA R16F2n/R16R2 fragment indicated that the phytoplasma associated with HFWB disease belongs to subgroup 16SrV-B of the elm yellows (EY) phytoplasma group. Nucleotide sequence analysis of the cloned HFWB phytoplasma partial rRNA operon (GenBank Accession No. EF029092), spanning a near full-length 16S rRNA gene and a partial 16S-23S rRNA intergenic spacer, suggested that HFWB phytoplasma is most closely related to JWB and PMWB phytoplasmas, both members of subgroup16SrV-B. To further characterize the HFWB phytoplasma, a genomic segment covering full-length ribosomal protein genes rplV and rpsC was PCR-amplified using primer pair rp(V)F1A/rp(V)R1A (2), cloned, and sequenced (GenBank Accession No. EF029093). The nucleotide sequence of the HFWB phytoplasma rplV and rpsC locus is nearly identical (99.9%) to that of JWB phytoplasma. To our knowledge, this is the first report of a phytoplasmal disease in Cannabis spp. Since HFWB and JWB phytoplasmas share extremely high sequence identity and share the same eco-geographic location, further investigation is warranted to determine whether these two phytoplasmas are actually one species that can infect both plants, an issue having important implications in managing both diseases. References: (1) S. Hong and R. C. Clarke. J. Int. Hemp Assoc. 3:55, 1996. (2) I. M. Lee et al. Int. J. Syst. Evol. Microbiol. 54:337, 2004. (3) Q. Liu et al. Plant Dis. 88:770, 2004. (4) Q. Liu et al. Plant Dis. 89:529, 2005.

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