scholarly journals Molecular characterisation of phytoplasmas infecting roses in Poland

2013 ◽  
Vol 55 (1) ◽  
pp. 325-334
Author(s):  
Hanna Śliwa ◽  
Tadeusz Malinowski ◽  
Maria Kamińska
Keyword(s):  
Reference Strain ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Nested Polymerase Chain Reaction ◽  
Aster Yellows ◽  
Leaf Chlorosis ◽  
Leaf Yellowing ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Aster Yellows Phytoplasma

Symptoms of shoot dieback and leaf yellowing followed by leaf chlorosis were observed in naturally infected roses 'Frisco' and 'Suela', cultivated in a commercial greenhouse in Poland. The presence of phytoplasma was demonstrated in affected plants by nested polymerase chain reaction (PCR) with R16Fl/RO and Pl/P7 primer pairs in the first round followed by a second one with R16F2n/R2, fA/rA, Pc399/P1694, R16(I)Fl/Rl and Pl/fArev primer pairs. Restriction fragment length polymorphism (RFLP) analysis of PCR products (primed with primers R16F2n/R2) was done using enzymes AluI, MseI, RsaI and HpaII. Restriction profiles obtained with these enzymes were identical to those of reference strain AY1 belonging to aster yellows phytoplasma group, subgroup I-B (16SrI-B). Nested PCR products from roses 'Frisco' and 'Suela' were sequenced. Analysis of sequences confirmed that the phytoplasma infecting those roses could be classified to aster yellows phytoplasma group, subgroup B.

scholarly journals First Report of Phytoplasma Belonging to Apple Proliferation Group in Roses in Poland

Plant Disease ◽  
2004 ◽  
Vol 88 (11) ◽  
pp. 1283-1283 ◽  
Author(s):  
M. Kamińska ◽  
H. Śliwa
Keyword(s):  
Reference Strain ◽  
Rflp Analysis ◽  
The United States ◽  
Apple Proliferation ◽  
Specific Primer ◽  
Disease Symptoms ◽  
Aster Yellows ◽  
Pcr Products ◽  
Aster Yellows Phytoplasma ◽  
Reference Strains

Virus-like diseases of rose plants of uncertain aetiology have been widely distributed throughout the world (4). Symptoms include dieback, rose rosette, witches' broom, and bud proliferation. Research conducted in England (4) and the United States (1) could not reveal the etiology of the diseases. Disease symptoms including stunted growth, leaf and flower malformation, and shoot and flower proliferation were observed in rose plants in Poland (2). In this previous work, we reported cases of phytoplasma closely related to group 16SrI in rose plants with shoot proliferation as determined by nested polymerase chain reaction (PCR) with ribosomal primers R16F1/R0 followed by rA/fA or R16(I)F1/R1. In this study, we examined 48 symptomatic rose plants of 12 cultivars using nested PCR primed by P1/P7 and followed by universal primer pairs R16F2n/R2, fA/rA, or group 16SrI-specific R16(I)F1/R1. To detect potential mixed infection in roses, the group 16SrX-specific primer pairs fAT/rAS, fAT/rPRUS, and fPD/rAT were used for nested PCRs. Samples of rose plants with disease symptoms and nonsymptomatic, samples of Catharanthus roseus, healthy and inoculated by grafting with the reference strain of aster yellows phytoplasma (AY1, group 16SrI-B, kindly supplied by I.-M. Lee, Beltsville, MD) and the reference strain of apple proliferation phytoplasma (AP, group 16SrX-A, kindly supplied by A. Bertaccini, Bologna, Italy), were tested for the presence of phytoplasma rDNA by PCR. Phytoplasma identification was accompanied by digestion with restriction endonucleases, AluI, HhaI, HpaII, MseI, and RsaI, and restriction fragment length polymorphism (RFLP) analysis of a 1.2-kb fragment of rDNA (3). A DNA amplification product was observed in all nested PCRs containing template DNA of samples collected from diseased roses and the reference strains but not from control plants. On the basis of RFLP analysis of PCR products and comparison of the RFLP patterns with those of the reference strains, we demonstrated the presence of aster yellows phytoplasma belonging to phytoplasma group 16SrI-B in roses of 11 cultivars. RFLP profile of samples collected from rose cv. Red Champ was identical to those obtained for reference AP strain (group 16SrX-A). Mixed RFLP profiles were observed in samples collected from rose cv. Memory, which were doubly infected by phytoplasmas belonging to groups 16SrI-B or 16SrX-A. These results were confirmed by PCR with group 16SrX-specific primer pairs. The target DNA was amplified when amplifications were conducted with subgroup 16SrX-A-specific primer pair fAt/rAS, whereas no observable PCR products were obtained with subgroup 16SrX-B- (fAT/rPRUS) or 16SrX-C- (fPD/rAT) specific primer pairs. This report confirms infection of roses by aster yellows phytoplasma belonging to group 16SrI-B, and to our knowledge, records for the first time, infection by phytoplasma of group 16SrX-A. References: (1) A. H. Epstein and J. H. Hill. J. Phytopathol. 143:353, 1995. (2) M. Kami ska et al. J. Phytopathol. 149:3, 2001. (3) I.-B. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1968. (4) B. J. Thomas. Ann. Rep. Glasshouse Crops Res. Inst. 1979:178, 1981.

scholarly journals First Report of “Candidatus Phytoplasma asteris”-Related Strains in Brassica rapa in Saskatchewan, Canada

Plant Disease ◽  
2006 ◽  
Vol 90 (6) ◽  
pp. 832-832 ◽  
Author(s):  
C. Y. Olivier ◽  
G. Séguin-Swartz ◽  
D. Hegedus ◽  
T. Barasubiye
Keyword(s):  
Sequence Data ◽  
Direct Sequencing ◽  
Nested Polymerase Chain Reaction ◽  
Aster Yellows ◽  
First Report ◽  
Leaf Yellowing ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Subgroup Member ◽  
Root Tissues

“Candidatus phytoplasma asteris” and related strains (i.e., aster yellows group 16SrI) have been associated with diseases of numerous plant species worldwide. Symptoms of aster yellows (AY) have been reported on rapeseed/canola (Brassica napus and B. rapa) crops in Saskatchewan (SK) and Manitoba, Canada since 1953 (2). Symptoms generally include stunting, virescence, leaf yellowing or purpling, phyllody, and formation of bladder-like siliques. A total of 120 mature B. rapa cv. AC Sunbeam plants exhibiting AY symptoms were collected in commercial fields near Medstead, SK during 2003 and 2004 (one field per year). As described previously (4), total genomic DNA was extracted from leaf, stem, roots, and seeds collected from the 120 plants, from seeds from the seed lots sown in 2003 and 2004, and from leaf and stem tissue of 20 greenhouse-grown plants from each seed lot. The latter DNA samples were assayed for phytoplasma DNA by a nested polymerase chain reaction (PCR) assay incorporating phytoplasma universal 16S rRNA primer pairs P1/P6 (1) followed by R16R2/R16F2 (4). Seed samples analyzed from the 2003 and 2004 seed lots and tissues of the 40 greenhouse-grown plants all tested negative for phytoplasma DNA using this assay. Leaf, stem, and/or root tissues of all plants collected in the field in 2003 (60 plants) and 2004 (60 plants) and 71.1% (315 of 443) of seed samples (five seeds per sample) tested positive for the presence of phytoplasma DNA, as evidenced by the presence of an expected band of 1.2 kb on the gels after the second amplification with primers R16R2/R16F2. Nested PCR products from plant samples collected in 2003 were cloned, sequenced, and compared with phytoplasma sequences archived in the GenBank nucleotide database. On this basis, phytoplasmas detected in plants or their seeds collected in 2003 were found to be most similar (98.8%) to CHRY (Accession No. AY180956), a 16SrI-A subgroup strain, or were most similar (98.9%) to isolate 99UW89 (Accession no. AF268407), a known 16SrI-B subgroup strain. Sequences of phytoplasmas detected in plants or their seeds in 2004 were obtained by direct sequencing of rRNA products amplified from samples using PCR incorporating primer pairs P1/P6 and P4/P7 (3). Analysis of sequence data revealed that phytoplasmas in these plants were all most similar (99.5%) to AY-WB (Accession no. AY389828), a 16SrI-A subgroup member. The nucleotide sequences have been deposited with GenBank under Accession nos. DQ404346, DQ404347, and DQ411470. To our knowledge, this is the first report of 16SrI-A and 16SrI-B subgroup phytoplasmas infecting plants and seed of B. rapa in Saskatchewan. References: (1) I.-M. Lee et al. Phytopathology, 83:834, 1993. (2) W. E. Sackston. Can. Plant Dis. Surv. 33:41, 1953. (3) L. B. Sharmila et al. J. Plant Biochem. Biotech. 13:1, 2004. (4) E. Tanne et al. Phytopathology, 91:741, 2001.

scholarly journals Detection and Characterization of a Lethal Yellowing (16SrIV) Group Phytoplasma in Canary Island Date Palms Affected by Lethal Decline in Texas

Plant Disease ◽  
2002 ◽  
Vol 86 (6) ◽  
pp. 676-681 ◽  
Author(s):  
N. A. Harrison ◽  
M. Womack ◽  
M. L. Carpio
Keyword(s):  
Canary Island ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Corpus Christi ◽  
Rdna Sequences ◽  
Leaf Yellowing ◽  
Pcr Products ◽  
Lethal Yellowing ◽  
Polymerase Chain

Polymerase chain reaction (PCR) assays were used to detect phytoplasmas in Canary Island date (Phoenix canariensis) palms displaying symptoms similar to lethal yellowing (LY) disease in Corpus Christi, TX. An rDNA product (1.8 kb) was amplified consistently from 10 of 11 palms by PCR employing phytoplasma universal rRNA primer pair P1/P7. Also, AluI endonu-clease digests and sequencing of P1/P7 products revealed that nontarget Bacillus megaterium-related rDNA sequences of similar size were co-amplified along with phytoplasma rDNA from 10 palms. A 1,402-bp product was obtained from all 11 symptomatic palms when initial P1/P7 products were reamplified by PCR employing nested LY phytoplasma group-specific 16S rRNA primer pair LY16Sf/LY16Sr. Restriction fragment length polymorphism (RFLP) analysis of nested PCR products revealed that palm-infecting phytoplasmas were uniform and most similar to strains composing the coconut lethal yellowing phytoplasma (16SrIV) group. Sequence analysis of 16S rDNA determined the Texas Phoenix palm decline (TPD) phytoplasma to be phylogenetically closest to the Carludovica palmata leaf yellowing (CPY) phytoplasma. rDNA profiles of strains TPD and CPY obtained with AluI were co-identical and distinct from other known 16SrIV group phytoplasmas. On this basis, both strains were classified as members of a new subgroup, 16SrIV-D.

scholarly journals Molecular Characterization and Classification of Phytoplasmas Associated with Canola Yellows and a New Phytoplasma Strain Associated with Dandelions

Plant Disease ◽  
2001 ◽  
Vol 85 (1) ◽  
pp. 76-79 ◽  
Author(s):  
Keri Wang ◽  
Chuji Hiruki
Keyword(s):  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Universal Primers ◽  
Dna Fragments ◽  
Chain Reaction ◽  
Aster Yellows ◽  
Distinct Subgroup ◽  
16S Ribosomal Dna ◽  
Polymerase Chain

DNA isolated from symptomatic canola (Brassica napus, Brassica rapa) and dandelion (Taraxacum officinale) was used to amplify 16S ribosomal DNA fragments by polymerase chain reaction using two pairs of universal primers P1/P6 and R16F2n/R2. Restriction fragment length polymorphism (RFLP) analysis of the amplified DNA fragments using endonucleases AluI, HhaI, HpaII, MseI, RsaI, and Sau 3AI revealed two distinct types of phytoplasmas in canola with similar symptoms. One had the same RFLP profiles as the phytoplasmas in subgroup 16SrI-A, whereas the other one had RFLP profiles similar to those of phytoplasmas in subgroup 16SrI-B. Phytoplasmas were detected in symptomatic dandelion plants that were collected from canola and alfalfa fields where severe alfalfa witches'-broom occurred. Comparative studies indicated that two different phytoplasmas were associated with the dandelion plants. One was identified as a member of subgroup 16SrI-A, whereas another one was classified as a member of a distinct subgroup in the aster yellows group on the basis of the unique RFLP patterns.

scholarly journals Molecular identification of Candidatus Phytoplasma spp. associated with Sophora yellow stunt in Iran

2017 ◽  
Vol 57 (2) ◽  
pp. 167-172 ◽  
Author(s):  
Touhid Allahverdi ◽  
Heshmatollah Rahimian ◽  
Mina Rastgou
Keyword(s):  
Phylogenetic Trees ◽  
Fragment Length Polymorphism ◽  
Chain Reaction ◽  
Mazandaran Province ◽  
Nested Polymerase Chain Reaction ◽  
Sophora Alopecuroides ◽  
Leaf Yellowing ◽  
Tehran Province ◽  
Polymerase Chain ◽  
West Azerbaijan

Abstract In the spring of 2012, sophora (Sophora alopecuroides L.) plants showing symptoms of leaf yellowing, little leaves and stunting were observed in Firooz-kuh (Tehran province), Sari (Mazandaran province) and Urmia (West Azerbaijan province) in Iran. Symptomatic plants from the three locations were subjected to nested polymerase chain reaction (PCR) to amplify 16SrRNA using primer pair P1/P7 followed by primer pair R16F2n/R16R2. The amplicons were purified, sequenced and the nucleotide sequences were analyzed by virtual restriction fragment length polymorphism (RFLP). The phytoplasmas associated with the yellows disease were identified as members of the 16SrIX group (Candidatus Phytoplasma phoenicium) and the 16SrXII group (Candidatus Phytoplasma solani). The two phytoplasmas were placed in 16SrIX-C and 16SrXII-A subgroups, respectively, in constructed phylogenetic trees. This is the first report on sophora yellows associated with Candidatus Phytoplasma phoenicium.

scholarly journals First Report of an Aster Yellows Subgroup 16SrI-B Phytoplasma Infecting Chayote in Costa Rica

Plant Disease ◽  
2002 ◽  
Vol 86 (3) ◽  
pp. 330-330 ◽  
Author(s):  
W. Villalobos ◽  
L. Moreira ◽  
C. Rivera ◽  
K. D. Bottner ◽  
I.-M. Lee
Keyword(s):  
Costa Rica ◽  
Rflp Analysis ◽  
Restriction Enzymes ◽  
Potential Threat ◽  
Nested Polymerase Chain Reaction ◽  
Aster Yellows ◽  
First Report ◽  
Rdna Sequences ◽  
Pcr Products ◽  
Production Area

An outbreak of a witches' broom disease affected approximately 20% of plants in several chayote (Sechium edule (Jacq.) Schwartz) fields in the commercial production area of the Ujarrás Valley, Cartago Province, Costa Rica during 2000 and 2001. Affected chayote plants exhibited symptoms, including basal proliferation with severe foliage reduction, aborted flowers, and deformed fruits, suggestive of phytoplasmal infection. Two other symptomatic cucurbit species growing near the chayote fields were also identified. These species were tacaco plants (S. tacaco (Pitt.) C. Jeffrey), an edible cucurbit for domestic marketing in Costa Rica, showing severe size reduction of leaves and fruits, and Rytidostylis carthaginensis (Jacq.) Kuntze, a weed in chayote and tacaco fields, exhibiting abnormal tendril proliferation. Plants were analyzed for phytoplasma infection by a nested polymerase chain reaction (PCR) assay, using the universal rRNA primer pair P1/P7 followed by R16F2n/R16R2 (2). Phytoplasmas were detected in all symptomatic samples (18 chayote, 6 tacaco, and 3 weed) tested but were undetectable in all asymptomatic samples (10 chayote, 6 tacaco, and 2 weed). Restriction fragment length polymorphism (RFLP) analysis of PCR products (16S rDNA sequences) by separate digestion with eight restriction enzymes (RsaI, HhaI, KpnI, BfaI, HaeIII, HpaII, AluI, MseI) revealed that a phytoplasma belonging to subgroup 16SrI-B in the aster yellows phytoplasma group (16SrI) was associated with chayote witches' broom (CWB). The same or very similar phytoplasmas were found in both symptomatic tacaco and R. carthaginensis plants. Phylogenetic analysis of 16SrDNA sequences also confirmed the CWB phytoplasma to be most similar to members of subgroup 16SrI-B. Similar diseases in chayote and other cucurbits have been reported in Brazil (3), Taiwan (1), and Mexico (4). The CWB phytoplasma differs from the phytoplasma (16SrIII-J subgroup) associated with chayote in Brazil. The identities of phytoplasmas associated with cucurbits in Taiwan and Mexico are unknown. The occurrence of an aster yellows group phytoplasma in chayote may pose a potential threat to continued production and exportation of this cash crop. To our knowledge, this is the first report of 16SrI-B subgroup phytoplasmas in naturally infected cucurbits in Costa Rica. References: (1) T. G. Chou et al. Plant Dis. Rep. 60:378, 1976. (2) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (3) H. G. Montano et al. Plant Dis. 84:429, 2000. (4) E. Olivas. Rev. Fitopatol. (Lima) 13:14, 1978.

scholarly journals ‘Candidatus Phytoplasma asteris’ Strains Associated with Oil Palm Lethal Wilt in Colombia

Plant Disease ◽  
2014 ◽  
Vol 98 (3) ◽  
pp. 311-318 ◽  
Author(s):  
Elizabeth Alvarez ◽  
Juan F. Mejía ◽  
Nicoletta Contaldo ◽  
Samanta Paltrinieri ◽  
Bojan Duduk ◽  
...  
Keyword(s):  
Oil Palm ◽  
Sequence Data ◽  
Severe Disease ◽  
Rflp Analysis ◽  
Nested Polymerase Chain Reaction ◽  
Aster Yellows ◽  
Rdna Sequences ◽  
Polymerase Chain ◽  
Reference Strains

The distribution of lethal wilt, a severe disease of oil palm, is spreading throughout South America. An incidence of about 30% was recorded in four commercial fields in Colombia. In this study, phytoplasmas were detected in symptomatic oil palm by using specific primers, based on 16S ribosomal DNA (rDNA) sequences, in nested polymerase chain reaction assays. The phytoplasmas were then identified as ‘Candidatus Phytoplasma asteris’, ribosomal subgroup 16SrI-B, through the use of restriction fragment length polymorphism (RFLP) analysis and sequencing. Cloning and sequencing of 16S rDNA from selected strains, together with phylogenetic analysis, confirmed the classification. Moreover, collective RFLP characterization of the groEL, amp, and rp genes, together with sequence data, distinguished the aster yellows strain detected in Colombian oil-palm samples from other aster yellows phytoplasmas used as reference strains; in particular, from an aster yellows strain infecting corn in the same country.

Identification of Borrelia burgdorferi genospecies inducing Lyme disease in dogs from Western Poland

2005 ◽  
Vol 53 (1) ◽  
pp. 13-21 ◽  
Author(s):  
Bogumiła Skotarczak ◽  
Beata Wodecka
Keyword(s):  
Borrelia Burgdorferi ◽  
Fragment Length Polymorphism ◽  
Sensu Stricto ◽  
Endemic Region ◽  
Nested Polymerase Chain Reaction ◽  
Veterinary Clinic ◽  
Reference Isolate ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Northwestern Poland

Canine Lyme borreliosis may be caused by three Borrelia burgdorferi sensu lato genospecies. The prevalence of infection by Borrelia species was determined by nested polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) with the enzyme Fsp4H I in the blood of dogs naturally infested by ticks in an endemic region of Poland. Blood samples were collected from 98 dogs of various breeds, delivered to the Veterinary Clinic in Szczecin (northwestern Poland) for various reasons. Nested PCR revealed the presence of DNA characteristic of only 1 genospecies, i.e. B. burgdorferi sensu stricto (s.s.), in all PCR-positive samples. Digestion of PCR products from a fragment of the fla gene amplified with primers FLA1 and FLA2 gave only one band pattern consistent with the pattern obtained from sequence analysis of the fla gene from a reference isolate of B. burgdorferi s.s. GeHo (X15660) from GenBank.

scholarly journals First Report of Alfalfa Witches Broom Disease in Oman Caused by a Phytoplasma of the 16Sr II Group

Plant Disease ◽  
2001 ◽  
Vol 85 (12) ◽  
pp. 1287-1287 ◽  
Author(s):  
A. J. Khan ◽  
K. M. Azam ◽  
M. L. Deadman ◽  
A. M. Al-Subhi ◽  
P. Jones
Keyword(s):  
Sweet Potato ◽  
Infected Plant ◽  
Rflp Analysis ◽  
Negative Control ◽  
Sultanate Of Oman ◽  
Nested Polymerase Chain Reaction ◽  
Aster Yellows ◽  
First Report ◽  
Pcr Products ◽  
Plant Pathol

Alfalfa (Medicago sativa L.) is a primary forage crop in the Sultanate of Oman. A new disease of alfalfa in Oman is characterized by proliferation of shoots and yellowing of leaves in 1- to 2-year-old plants and tillering of stems in 4- to 5-year-old plants. Annual losses due to this disease are estimated at more than US$ 23 million. Samples of healthy and infected alfalfa plants were collected from different regions. Total DNA was extracted according to Khadhair et al. (1), with minor modifications. Amplification of 16S rDNA was done using a nested polymerase chain reaction (PCR) approach with primers P1/P7 and R16F2n/R16R2. DNA from healthy leaves and sterile water was used as a negative control, while DNA from periwinkle infected with faba bean phyllody (16SrII-C), aster yellows (16SrI), tomato big bud (16SrII-D), sweet potato little leaf (16SrII-D), catharanthus phyllody (16SrVI), and sesame phyllody (16SrII-A) were used as positive controls and for restriction fragment length polymorphism (RFLP) comparisons. Nested 1.25-kb PCR products from infected plant samples were subjected to RFLP analysis with restriction endonucleases RsaI, AluI, HaeIII, HhaI, EcoRI, TaqI, Tru9I, and Sau3AI. The analysis showed that the alfalfa witches' broom phytoplasma (AWBP) belonged to the 16SrII group (peanut witches' broom) and that the AWBP was most similar to sweet potato little leaf (16SrII-D) but distinct from “Candidatus Phytoplasma aurantifolia,” the cause of lime witches' broom in Oman. Other phytoplasmas infecting alfalfa have been reported from Europe and North America (1,3), but they belong to the 16SrVI (clover phyllody) and 16SrI (aster yellows) groups. An alfalfa witches' broom reported from Italy (2) forms a separate grouping (4). To our knowledge, this is the first report of a phytoplasma from the peanut witches' broom group infecting alfalfa in the Sultanate of Oman. References: (1) A. H. Khadhair et al. Microbiol. Res. 152:259, 1997. (2) C. Marcone et al. J. Plant Pathol. 79:211, 1997. (3) R. D. Peters et al. Plant Dis. 83:488, 1999. (4) E. Seemuller et al. J. Plant Pathol. 80:3, 1998.

scholarly journals First Report of a Group 16SrI, Subgroup B, Phytoplasma in Diseased Epilobium hirsutum in the Region of Tallin, Estonia

Plant Disease ◽  
2002 ◽  
Vol 86 (10) ◽  
pp. 1177-1177 ◽  
Author(s):  
A. Alminaite ◽  
R. E. Davis ◽  
D. Valiunas ◽  
R. Jomantiene
Keyword(s):  
16S Rdna ◽  
Sequence Similarity ◽  
Intergenic Spacer ◽  
Rflp Analysis ◽  
Nested Polymerase Chain Reaction ◽  
Universal Primer ◽  
Aster Yellows ◽  
First Report ◽  
Intergenic Spacer Region ◽  
Aster Yellows Phytoplasma

Symptoms of phyllody of flowers and general plant yellowing indicating possible phytoplasma infection were observed in diseased plants of hairy willow-weed (Epilobium hirsutum L., family Onagraceae) growing in a meadow at Harku Village near Tallin, Estonia. DNA was extracted from diseased E. hirsutum using a Genomic DNA Purification Kit (Fermentas AB, Vilnius, Lithuania) and used as a template in nested polymerase chain reaction (PCR). Ribosomal (r) DNA was initially amplified in PCR primed by phytoplasma universal primer pair P1/P7 (4) and reamplified in PCR primed by nested primer pair 16SF2n/16SR2 (F2n/R2) (1) as previously described (2). Products of 1.8 kbp and 1.2 kbp were obtained in PCR primed P1/P7 and F2n/R2, respectively, from all four symptomatic plants examined. These data indicated that the diseased E. hirsutum plants were infected by a phytoplasma, termed epilobium phyllody (EpPh) phytoplasma. The 16S rDNA amplified in PCR primed by nested primer pair F2n/R2 was subjected to restriction fragment length polymorphism (RFLP) analysis using restriction endonucleases AluI, MseI, HpaI, HpaII, HhaI, RsaI, HinfI, and HaeIII (Fermentas AB). On the basis of the collective RFLP profiles, EpPh phytoplasma was classified in group 16SrI (aster yellows phytoplasma group), subgroup B (aster yellows phytoplasma subgroup), according to the phytoplasma classification scheme of Lee et al. (3). The 1.8-kbp rDNA product of P1/P7-primed PCR, which included 16S rDNA, 16S-23S intergenic spacer region, and the 5′ -end of 23S rDNA, was cloned in Escherichia coli using the TOPO TA Cloning Kit (Invitrogen, Carlsbad, Ca) according to manufacturer's instructions and sequenced. The sequence was deposited in the GenBank database as Accession No. AY101386. This nucleotide sequence shared 99.8% sequence similarity with a comparable rDNA sequence (GenBank Accession No. AF322644) of aster yellows phytoplasma AY1, a known subgroup 16SrI-B strain. The EpPh phytoplasma sequence was highly similar (99.9%) to operons rrnA (GenBank Accession No. AY102274) and rrnB (GenBank Accession No. AY102273) from Valeriana yellows (ValY) phytoplasma infecting Valeriana officinalis plants in Lithuania. ValY phytoplasma was found to exhibit rRNA interoperon sequence heterogeneity (D. Valiunas, unpublished data). To our knowledge, this is the first report to reveal E. hirsutum as a host of phytoplasma and to demonstrate the occurrence of a plant pathogenic mollicute in the northern Baltic region. References: (1) D. E. Gundersen and I.-M. Lee. Phytopathol. Mediterr. 35:144, 1996. (2) R. Jomantiene et al. HortScience 33:1069, 1998. (3) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (4) B. Schneider et al. Phlogenetic classification of plant pathogenic mycoplasma-like organisms or phytoplasmas. Page 369 in: Molecular and Diagnostic Procedures in Mycoplasmology, Vol 1, R. Razin, and J. G. Tully eds. Academic Press, San Diego, 1995.

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