Functional analysis of a lipolytic protein encoded in phytoplasma phage based genomic island.

Wall-less bacteria known as phytoplasmas are obligate transkingdom parasites and pathogens of plants and insect vectors. These unusual bacteria possess some of the smallest genomes known among pathogenic bacteria, and have never been successfully isolated in artificial culture. Disease symptoms induced by phytoplasmas in infected plants include abnormal growth and often severe yellowing of leaves, but mechanisms involved in phytoplasma parasitism and pathogenicity are little understood.

Evaluation of the DNA-dependent RNA polymerase β-subunit gene (rpoB) for phytoplasma classification and phylogeny.

Phytoplasmas are classified into 16Sr groups and subgroups and 'Candidatus Phytoplasma' species, largely or entirely based on analysis of 16S rRNA gene sequences. Yet, distinctions among closely related 'Ca. Phytoplasma' species and strains based on 16S rRNA genes alone have limitations imposed by the high degree of rRNA nucleotide sequence conservation across diverse phytoplasma lineages and by the presence in a phytoplasma genome of two, sometimes sequence-heterogeneous, copies of the 16S rRNA gene.

'Candidatus Phytoplasma pruni', a novel taxon associated with X-disease of stone fruits, Prunus spp.: multilocus characterization based on 16S rRNA, secY, and ribosomal protein genes.

X-disease is one of the most serious diseases known in peach (Prunus persica). Based on RFLP analysis of 16S rRNA gene sequences, peach X-disease phytoplasma strains from eastern and western United States and eastern Canada were classified in 16S rRNA gene RFLP group 16SrIII, subgroup A. Phylogenetic analyses of 16S rRNA gene sequences revealed that the X-disease phytoplasma strains formed a distinct subclade within the phytoplasma clade, supporting the hypothesis that they represented a lineage distinct from those of previously described 'Candidatus Phytoplasma' species.

'Candidatus Phytoplasma sudamericanum', a novel taxon, and strain PassWB-Br4, a new subgroup 16SrIII-V phytoplasma, from diseased passion fruit (Passiflora edulis f. flavicarpa Deg.).

Symptoms of abnormal proliferation of shoots resulting in formation of witches'-broom growths were observed on diseased plants of passion fruit (Passiflora edulis f. flavicarpa Deg.) in Brazil.

First Report of a New Phytoplasma Subgroup, 16SrIII-T, Associated with Decline Disease Affecting Sweet and Sour Cherry Trees in Lithuania.

During July 2007, sweet (Prunus avium) and sour cherry (P. cerasus) trees exhibiting disease symptoms suggestive of possible phytoplasma infection were observed in a large orchard in the Kaunas Region of Lithuania. Samples of leaf tissue were collected from 13 sweet cherry trees that were affected by a decline disease (designated cherry decline, ChD) characterized by symptoms that included leaf reddening and premature leaf drop and two sour cherry trees exhibiting proliferation of branches and nonseasonal flowering.

Ancient, recurrent phage attacks and recombination shaped dynamic sequence-variable mosaics at the root of phytoplasma genome evolution.

Mobile genetic elements have impacted biological evolution across all studied organisms, but evidence for a role in evolutionary emergence of an entire phylogenetic clade has not been forthcoming. We suggest that mobile element predation played a formative role in emergence of the phytoplasma clade. Phytoplasmas are cell wall-less bacteria that cause numerous diseases in plants.

Sequence-variable mosaics: composites of recurrent transposition characterizing the genomes of phylogenetically diverse phytoplasmas.

Phytoplasmas are cell wall-less prokaryotes characterized by small, AT-rich genomes that encode capabilities for obligate, transkingdom parasitism and pathogenicity in plants and insect vectors. Inability to isolate and characterize phytoplasmas in pure culture has led to adoption of the 'Candidatus species' convention to refer to distinct phytoplasma lineages. In this study, we provide evidence that multiple, sequence-variable mosaics (SVMs) of clustered genes and repetitive extragenic palindromes are characteristic features of phytoplasma genome architecture in phylogenetically diverse species.

Clusters of diverse genes existing as multiple, sequence-variable mosaics in a phytoplasma genome.

Phytoplasmas are cell wall-less prokaryotes living as obligate parasites and pathogens of plants and insects, making them attractive subjects for studies to gain a greater understanding of transkingdom parasitism and pathogenicity. During a study of two phytoplasma genomes, we obtained evidence for previously unreported clustering of genes, pseudogenes, mobile genetic elements, intergenic repeat units, and repetitive extragenic palindromes that occur in multiple, homologous clusters in some phytoplasma genomes. The clusters represent previously unrecognized mosaics, possibly assembled through multiple events of targeted mobile element attack, duplication, recombination, and rearrangement.

Lineage-specific decay of folate biosynthesis genes suggests ongoing host adaptation in phytoplasmas.

Phytoplasmas are nonculturable cell wall-less, obligate intracellular pathogens of plants and insect vectors. In their descent from walled bacterial ancestors, phytoplasmas underwent massive genome reduction, resulting in some of the smallest cellular genomes known in nonsymbiotic bacteria. While requirements for in vitro culture of phytoplasmas remain unknown, two opposing reports have appeared concerning genes encoding the ability of phytoplasmas to synthesize folates de novo.

Cryptic plasmid pSKU146 from the wall-less plant pathogen Spiroplasma kunkelii encodes an adhesin and components of a type IV translocation-related conjugation system.

A cryptic plasmid of the wall-less plant pathogenic mollicute, Spiroplasma kunkelii CR2-3X, was cloned and its sequence analyzed. The 14,615 bp plasmid, designated pSKU146, has a nucleotide content of 28 mol% G + C, and contains 18 potential protein-coding regions (open reading frames, ORFs), of which six encode proteins that exhibit similarity to virulence-associated proteins involved in cell-to-cell adhesion or conjugal DNA transfer. One ORF encodes a 96 kDa protein, SkARP1, that is highly similar to SARP1 adhesin involved in attachment of Spiroplasma citri to insect vector gut membrane.

First Report of Barley as Host of a Phytoplasma Belonging to Group 16SrI, Subgroup B, and Ribosomal Protein Subgroup rpI-B in Lithuania.

Numerous diseased plants of barley (Hordeum vulgaris L.) exhibiting twisted, abnormally thin and yellowed awns, reduced spikelets, and general stunting and yellowing were observed in fields in the Vilnius and Kaisiadorys regions of Lithuania. The possible association of a phyto-plasma with the disease, termed barley deformation (BaDef), was assessed using polymerase chain reaction (PCR).

Predicted ATP-binding cassette systems in the phytopathogenic mollicute Spiroplasma kunkelii.

Spiroplasma kunkelii is a cell wall-free, helical, and motile mycoplasma-like organism that causes corn stunt disease in maize. The bacterium has a compact genome with a gene set approaching the minimal complement necessary for cellular life and pathogenesis. A set of 21 ATP-binding cassette (ABC) domains was identified during the annotation of a draft S.

Folate biosynthesis pseudogenes, PsifolP and PsifolK, and an O-sialoglycoprotein endopeptidase gene homolog in the phytoplasma genome.

Phytoplasmas are wall-less phytopathogenic prokaryotes of small genome sizes that are obligate parasites of insect vectors and plant hosts. We have cloned a clover phyllody (CPh) phytoplasma DNA locus containing five potential coding sequences. Two were identified as pseudogenes (PsifolP and PsifolK) homologous to folP and folK genes, which encode dihydropteroate synthase (DHPS) and 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK), respectively, in other bacteria.

Differential amplification of sequence heterogeneous ribosomal RNA genes and classification of the 'Fragaria multicipita' phytoplasma.

Ribosomal (r) RNA interoperon sequence heterogeneity in the 'Fragaria multicipita' phytoplasma, a member of group 16SrVI, was initially observed in RFLP patterns of rDNA amplified in the polymerase chain reaction (PCR), and was confirmed through sequence analysis of cloned rDNA. Sequences from operons rrnA and rrnB were amplified in PCR primed by primer pair P1/P7 but from only rrnA in PCR primed by primer pair R16mF2/R16mR1. Preferential amplification of DNA from operon rrnA was explained by base mismatches between the R16mF2/R16mR1 primers and primer annealing sites in rrnB.

Gene content and organization of an 85-kb DNA segment from the genome of the phytopathogenic mollicute Spiroplasma kunkelii.

Spiroplasma kunkelii, the causative agent of corn stunt disease in maize (Zea maysL.), is a helical, cell wall-less prokaryote assigned to the class Mollicutes. As part of a project to sequence the entire S.

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

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.

Molecular Identification and Classification of Strawberry Phylloid Fruit Phytoplasma in Group 16SrI, New Subgroup.

Plants of commercial strawberry (Fragaria × ananassa Duch., cv. Camarosa) exhibiting extensive fruit phyllody (development of leafy structures from achenes) were observed in a winter greenhouse production facility in West Virginia.

First Report of Oat as Host of a Phytoplasma Belonging to Group 16SrI, Subgroup A.

Diseased plants of oat (Avena sativa L.) exhibiting abnormal proliferation of spikelets were observed in the field in Raseniai, Lithuania. The possible association of a phytoplasma with the disease, termed oat proliferation (OatP), was determined using polymerase chain reaction (PCR) for amplification of phytoplasmal ribosomal (r) RNA gene (rDNA) sequences from template DNA extracted from the diseased oats.

First Report of Alder Yellows Phytoplasma in the Eastern Baltic Region.

Alnus glutinosa (alder) is widespread in Europe and is an important component of biological diversity in natural forest ecosystems in the Baltic Region. In 2000, diseased trees of A. glutinosa exhibiting characteristically phytoplasmal disease symptoms of shoot proliferation and leaf yellowing were observed in Aukstaitija National Park, Lithuania.

First Report of Aster Yellows-Related Subgroup I-A Phytoplasma Strains in Carrot, Phlox, Sea-Lavender, Aconitum, and Hyacinth in Lithuania.

Phytoplasma strains that belong to group 16SrI (aster yellows phytoplasma group), subgroup A (I-A, North American tomato big bud phytoplasma subgroup) were discovered in diverse plant species in Lithuania. Plants in which the strains were found exhibited symptoms characteristic of infections by phytoplasma. Carrot (Daucus sativus) with carrot proliferation disease exhibited symptoms of proliferation of the crown, chlorosis of young leaves, and reddening of mature leaves.

Molecular Identification and Classification of a Phytoplasma Associated with Phyllody of Strawberry Fruit in Maryland.

Several phytoplasmas have been reported to be associated with phyllody of strawberry fruit, including clover yellow edge, clover proliferation, clover phyllody, eastern and western aster yellows, STRAWB2, strawberry multicipita, and Mexican periwinkle virescence phytoplasmas. Plant symptoms in addition to phyllody may include chlorosis, virescence, stunting, or crown proliferation. In this report we describe a new phytoplasma in association with strawberry leafy fruit (SLF) disease in Maryland.

Single and Mixed Phytoplasma Infections in Phyllody- and Dwarf-Diseased Clover Plants in Lithuania.

Naturally diseased plants of clover (Trifolium spp.) exhibiting symptoms of clover phyllody (virescence and phyllody of flowers) or of clover dwarf (abnormally small leaves, shortened internodes, proliferation of shoots, and dwarf growth habit) were observed in fields in Lithuania. Phytoplasma group-specific polymerase chain reactions (PCRs) and restriction fragment length polymorphism (RFLP) analysis of 16S rDNA revealed that the plants were infected by two mutually distinct phytoplasmas.

First Report of Phytoplasmas in Soybean, Alfalfa, and Lupinus sp. in Lithuania.

Plants of cultivated soybean (Glycine max) and alfalfa (Medicago sativa) in Dotnuva and of wild Lupinus sp. in Ledakalnis, Lithuania, exhibited symptoms that suggested phytoplasmal infections. Soybean plants were of normal growth habit but exhibited veinal necrosis.

First Report of Clover Yellow Edge Phytoplasma in Corylus (Hazelnut).

During investigations into the cause of a stunt syndrome affecting cultivated European hazelnut trees (Corylus avellana L.) in Oregon, the clover yellow edge (CYE) phytoplasma was detected for the first time in this crop. The cause of hazelnut stunt syndrome (HSS) is unknown, but the disease has been transmitted by grafting and apparently has moved within orchards through root grafts (1).

First Report of Clover Yellow Edge and STRAWB2 Phytoplasmas in Strawberry in Maryland.

Commercial strawberry (Fragaria × ananassa Duchesne) plants that were either chlorotic and severely stunted or exhibiting fruit phyllody were collected in Maryland. The plants were assessed for phytoplasma infection by nested polymerase chain reactions primed by phytoplasma universal primer pairs R16mF2/R1 and F2n/R2 (2) or P1/P7 (3) and F2n/R2 for amplification of phytoplasma 16S ribosomal (r) DNA (16S rRNA gene) sequences. Phytoplasma-characteristic 1.