New Disease Reports (2010) 20, 35.

First report of a new subgroup 16SrIX-E (‘Candidatus Phytoplasma phoenicium’-related) phytoplasma associated with juniper witches’ broom disease in Oregon, USA

R.E. Davis1*, E.L. Dally1, Y. Zhao1, I.-M. Lee1, R. Jomantiene2, A.J. Detweiler3 and M.L. Putnam4

1 USDA-Agricultural Research Service, Beltsville, MD 20705, USA
2 Institute of Botany, Vilnius, LT-08406, Lithuania
3 Oregon State University Deschutes County Extension, Redmond, OR 97756, USA
4 Oregon State University Plant Clinic, Corvallis, OR 97331-2903, USA

*robert.davis@ars.usda.gov

Accepted: 13 Jan 2010

Western juniper(Juniperus occidentalis) is a native tree indigenous to parts of Oregon, Washington, Idaho, Nevada and California (USA).The tree has increased in density since settlement of these areas, raising concern over loss of understory plants, decreased wildlife habitat and increased soil erosion.A newly recognized disease, juniper witches’ broom (JunWB), affecting at least 1% of trees in central Oregon, is characterized by abnormal proliferation of shoots, reduced size of leaves, shortened internodes, and growths having a ball-like appearance (Fig. 1).DNA was extracted from leaf samples from ball-like growths and used as template in polymerase chain reactions primed by primer pair P1/P7 (Deng & Hiruki, 1991; Schneider et al., 1995).DNA fragments of 1.8 kb amplified from two samples were sequenced and the sequences deposited in GenBank (Accession Nos. GQ925918 and GQ925919).RFLP patterns of 16S rDNA, observed as virtual patterns (Fig. 2) using iPhyClassifier (Zhao et al., 2009), indicated that JunWB phytoplasma represents a new subgroup lineage, designated 16SrIX-E.16S rDNA sequence similarity confirmed that JunWB is a ‘Ca. Phytoplasma phoenicium’-related phytoplasma. 

JunWB is one of two phytoplasmas found thus far to infect gymnosperms and is the only phytoplasma known to infect Juniperus sp.Occurrence of two distinct phytoplasmas, JunWB phytoplasma (this study) and ‘Ca. Phytoplasma pini’ (Schneider et al., 2005), in gymnosperms of two different families (Pinaceae and Cupressaceae, Division Coniferae) in Europe and North America, suggests that phytoplasmal infection of conifers may be more common than previously envisioned.

Figure1
Figure 1: Figure 1: Symptoms characteristic of juniper witches’-broom (JunWB) disease include abnormal proliferation of shoots, reduced size of leaves, shortened internodes and growths having a ball-like appearance
Figure2
Figure 2: Figure 2: Virtual RFLP patterns derived from in silico digestions of 1.2 kb 16S rDNA fragments. (A) Patterns from JunWB phytoplasma; digestion using 17 restriction endonuclease enzymes; (B) and (C) differentiation of JunWB from phytoplasmas belonging to previously designated subgroups in group 16SrIX by digestion of 16S rDNA with (B) AluI and (C) TaqI. Restriction fragments were resolved by in silico electrophoresis through 3% agarose gel. MW, ΦX174 DNA-HaeIII digestion.

References

Deng S, Hiruki C, 1991. Amplification of 16S rRNA genes from culturable and non-culturable mollicutes.Journal ofMicrobiological Methods 14, 53-61.

Schneider B., Seemüller E, Smart CD, Kirkpatrick BC, 1995. Phylogenetic classification of plant pathogenic mycoplasma-like organisms or phytoplasmas.In: Razin S, Tully JG, eds. Molecular and Diagnostic Procedures in Mycoplasmology. New York, USA: Academic Press, Vol. 2, 369-80.

Schneider B, Torres E, Martin MP, Schröder M, Behnke H-D, Seemüller E,2005. 'Candidatus Phytoplasma pini', a novel taxon from Pinus silvestris and Pinus halepensis.InternationalJournal ofSystematic andEvolutionary Microbiology 55, 303-307.

Zhao Y, Wei W, Lee I-M, Shao J, Suo X, Davis RE,2009. Construction of an interactive online phytoplasma classification tool, iPhyClassifier, and its application in analysis of the peach X-disease phytoplasma group (16SrIII).InternationalJournal ofSystematic andEvolutionary Microbiology59, 2582-2593.

©2010 The Authors