The trivial name ‘phytoplasma’ has been adopted to collectively name wall-less, non-helical prokaryotes that colonize plant phloem and insects, which were formerly known as mycoplasma-like organisms. Although phytoplasmas have not yet been cultivated in vitro, phylogenetic analyses based on various conserved genes have shown that they represent a distinct, monophyletic clade within the class Mollicutes. It is proposed here to accommodate phytoplasmas within the novel genus ‘Candidatus (Ca.) Phytoplasma’. Given the diversity within ‘Ca. Phytoplasma’, several subtaxa are needed to accommodate organisms that share <97·5 % similarity among their 16S rRNA gene sequences. This report describes the properties of ‘Ca. Phytoplasma’, a taxon that includes the species ‘Ca. Phytoplasma aurantifolia’ (the prokaryote associated with witches'-broom disease of small-fruited acid lime), ‘Ca. Phytoplasma australiense’ (associated with Australian grapevine yellows), ‘Ca. Phytoplasma fraxini’ (associated with ash yellows), ‘Ca. Phytoplasma japonicum’ (associated with Japanese hydrangea phyllody), ‘Ca. Phytoplasma brasiliense’ (associated with hibiscus witches'-broom in Brazil), ‘Ca. Phytoplasma castaneae’ (associated with chestnut witches'-broom in Korea), ‘Ca. Phytoplasma asteris' (associated with aster yellows), ‘Ca. Phytoplasma mali’ (associated with apple proliferation), ‘Ca. Phytoplasma phoenicium’ (associated with almond lethal disease), ‘Ca. Phytoplasma trifolii’ (associated with clover proliferation), ‘Ca. Phytoplasma cynodontis' (associated with Bermuda grass white leaf), ‘Ca. Phytoplasma ziziphi’ (associated with jujube witches'-broom), ‘Ca. Phytoplasma oryzae’ (associated with rice yellow dwarf) and six species-level taxa for which the Candidatus species designation has not yet been formally proposed (for the phytoplasmas associated with X-disease of peach, grapevine flavescence dorée, Central American coconut lethal yellows, Tanzanian lethal decline of coconut, Nigerian lethal decline of coconut and loofah witches'-broom, respectively). Additional species are needed to accommodate organisms that, despite their 16S rRNA gene sequence being >97·5 % similar to those of other ‘Ca. Phytoplasma’ species, are characterized by distinctive biological, phytopathological and genetic properties. These include ‘Ca. Phytoplasma pyri’ (associated with pear decline), ‘Ca. Phytoplasma prunorum’ (associated with European stone fruit yellows), ‘Ca. Phytoplasma spartii’ (associated with spartium witches'-broom), ‘Ca. Phytoplasma rhamni’ (associated with buckthorn witches'-broom), ‘Ca. Phytoplasma allocasuarinae’ (associated with allocasuarina yellows), ‘Ca. Phytoplasma ulmi’ (associated with elm yellows) and an additional taxon for the stolbur phytoplasma. Conversely, some organisms, despite their 16S rRNA gene sequence being <97·5 % similar to that of any other ‘Ca. Phytoplasma’ species, are not presently described as Candidatus species, due to their poor overall characterization.
Aster yellows (AY) group (16SrI) phytoplasmas are associated with over 100 economically important diseases worldwide and represent the most diverse and widespread phytoplasma group. Strains that belong to the AY group form a phylogenetically discrete subclade within the phytoplasma clade and are related most closely to the stolbur phytoplasma subclade, based on analysis of 16S rRNA gene sequences. AY subclade strains are related more closely to their culturable relatives, Acholeplasma spp., than any other phytoplasmas known. Within the AY subclade, six distinct phylogenetic lineages were revealed. Congruent phylogenies obtained by analyses of tuf gene and ribosomal protein (rp) operon gene sequences further resolved the diversity among AY group phytoplasmas. Distinct phylogenetic lineages were identified by RFLP analysis of 16S rRNA, tuf or rp gene sequences. Ten subgroups were differentiated, based on analysis of rp gene sequences. It is proposed that AY group phytoplasmas represent at least one novel taxon. Strain OAY, which is a member of subgroups 16SrI-B, rpI-B and tufI-B and is associated with evening primrose (Oenothera hookeri) virescence in Michigan, USA, was selected as the reference strain for the novel taxon ‘Candidatus Phytoplasma asteris’. A comprehensive database of diverse AY phytoplasma strains and their geographical distribution is presented.
Apple proliferation (AP), pear decline (PD) and European stone fruit yellows (ESFY) are among the most economically important plant diseases that are caused by phytoplasmas. Phylogenetic analyses revealed that the 16S rDNA sequences of strains of each of these pathogens were identical or nearly identical. Differences between the three phytoplasmas ranged from 1·0 to 1·5 % of nucleotide positions and were thus below the recommended threshold of 2·5 % for assigning species rank to phytoplasmas under the provisional status ‘Candidatus’. However, supporting data for distinguishing the AP, PD and ESFY agents at the species level were obtained by examining other molecular markers, including the 16S–23S rDNA spacer region, protein-encoding genes and randomly cloned DNA fragments. The three phytoplasmas also differed in serological comparisons and showed clear differences in vector transmission and host-range specificity. From these results, it can be concluded that the AP, PD and ESFY phytoplasmas are coherent but discrete taxa that can be distinguished at the putative species level, for which the names ‘Candidatus Phytoplasma mali’, ‘Candidatus Phytoplasma pyri’ and ‘Candidatus Phytoplasma prunorum’, respectively, are proposed. Strains AP15R, PD1R and ESFY-G1R were selected as reference strains. Examination of available data on the peach yellow leaf roll (PYLR) phytoplasma, which clusters with the AP, PD and ESFY agents, confirmed previous results showing that it is related most closely to the PD pathogen. The two phytoplasmas share 99·6 % 16S rDNA sequence similarity. Significant differences were only observed in the sequence of a gene that encodes an immunodominant membrane protein. Until more information on this phytoplasma is available, it is proposed that the PYLR phytoplasma should be regarded as a subtype of ‘Candidatus Phytoplasma pyri’.
Spartium witches'-broom (SpaWB), buckthorn witches'-broom (BWB) and allocasuarina yellows (AlloY) are witches'-broom and yellows diseases of Spartium junceum (Spanish broom), Rhamnus catharticus (buckthorn) and Allocasuarina muelleriana (Slaty she-oak), respectively. These diseases are associated with distinct phytoplasmas. The SpaWB, BWB and AlloY phytoplasmas share <97·5 % 16S rDNA sequence similarity with each other and with other known phytoplasmas, including the closely related phytoplasmas of the apple proliferation group. Also, the SpaWB, BWB and AlloY phytoplasmas each have a different natural plant host. Based on their unique properties, it is proposed to designate the mentioned phytoplasmas as novel ‘Candidatus’ species under the names ‘Candidatus Phytoplasma spartii’, ‘Candidatus Phytoplasma rhamni’ and ‘Candidatus Phytoplasma allocasuarinae’, respectively.
Bermuda grass white leaf (BGWL) is a destructive, phytoplasmal disease of Bermuda grass (Cynodon dactylon). The causal pathogen, the BGWL agent, differs from other phytoplasmas that cluster in the same major branch of the phytoplasma phylogenetic clade in <2·5 % of 16S rDNA nucleotide positions, the threshold for assigning species rank to phytoplasmas under the provisional status ‘Candidatus’. Thus, the objective of this work was to examine homogeneity of BGWL isolates and to determine whether there are, in addition to 16S rDNA, other markers that support delineation of the BGWL agent at the putative species level. Phylogenetic analyses revealed that the 16S rDNA sequences of BGWL strains were identical or nearly identical. Clear differences that support separation of the BGWL agent from related phytoplasmas were observed within the 16S–23S rDNA spacer sequence, by serological comparisons, in vector transmission and in host-range specificity. From these results, it can be concluded that the BGWL phytoplasma is a discrete taxon at the putative species level, for which the name ‘Candidatus Phytoplasma cynodontis' is proposed. Strain BGWL-C1 was selected as the reference strain. Phytoplasmas that are associated with brachiaria white leaf, carpet grass white leaf and diseases of date palms showed 16S rDNA and/or 16S–23S rDNA spacer sequences that were identical or nearly identical to those of the BGWL phytoplasmas. However, the data available do not seem to be sufficient for a proper taxonomic assignment of these phytoplasmas.
Previously, analysis of 16S rDNA sequences placed a newly discovered lineage of bacterial symbionts of arthropods in the ‘Bacteroidetes’. This symbiont lineage is associated with a number of diverse host reproductive manipulations, including induction of parthenogenesis in severalEncarsiaparasitoid wasps (Hymenoptera: Aphelinidae). In this study, electron microscopy and phylogenetic analysis of the 16S rRNA andgyrBgenes of symbionts fromEncarsia hispidaandEncarsia pergandiellaare used to describe and further characterize these bacteria. Phylogenetic analyses based on these two genes showed that theEncarsiasymbionts are allied with theCytophaga aurantiacalineage within the ‘Bacteroidetes’, with their closest described relative being the acanthamoeba symbiont ‘CandidatusAmoebophilus asiaticus’. TheEncarsiasymbionts share 97 % 16S rDNA sequence similarity withBrevipalpusmite andIxodestick symbionts and 88 % sequence similarity with ‘CandidatusA. asiaticus’. Electron microscopy revealed that many of the bacteria found in the ovaries of the twoEncarsiaspecies contained a regular, brush-like array of microfilament-like structures that appear to be characteristic of the symbiont. Finally, the role of this bacterium in parthenogenesis induction inE. hispidawas confirmed. Based on phylogenetic analyses and electron microscopy, classification of the symbionts fromEncarsiaas ‘CandidatusCardinium hertigii’ is proposed.
Elm yellows group (16SrV) phytoplasmas, which are associated with devastating diseases in elm, grapevine, blackberry, cherry, peach and several other plant species in America, Europe and Asia, represent one of the most diverse phytoplasma clusters. On the basis of phylogenetic analysis of 16S rDNA sequences, elm yellows group phytoplasmas form a discrete subclade within the phytoplasma clade. Three phylogenetic parameters, namely 16S rRNA, ribosomal protein and secY genes, have been evaluated for their usefulness in differentiating elm yellows group phytoplasmas. RFLP analysis of 16S rRNA sequences differentiated the elm yellows group phytoplasmas into five subgroups. Twelve RFLP subgroups were differentiated on the basis of ribosomal protein and 13 were differentiated using secY gene sequences. Phylogenetic analysis of the ribosomal protein genes and secY gene alone or in combination indicated that the subgroups constitute 12 genetically distinct lineages, each of which appears to have evolved under different ecological constraints such as specific vector or plant hosts. On the basis of unique DNA and biological properties, it is proposed that the elm yellows phytoplasma EY1T represents a novel taxon, ‘Candidatus Phytoplasma ulmi’.