The complete genome of "Candidatus Phytoplasma fraxini" AshY1 from the ash yellows group.

The complete genome of "Candidatus Phytoplasma fraxini" AshY1, originating from Fraxinus americana in North America, was assembled using long reads from single-molecule real-time sequencing technology. The chromosome of 598 kb provides insights into the effector repertoire of a phytopathogenic bacterium from the 16SrVII phytoplasma group.

Divergence within the Taxon 'Candidatus Phytoplasma asteris' Confirmed by Comparative Genome Analysis of Carrot Strains.

Phytoplasmas are linked to diseases in hundreds of economically important crops, including carrots. In carrots, phytoplasmosis is associated with leaf chlorosis and necrosis, coupled with inhibited root system development, ultimately leading to significant economic losses. During a field study conducted in Baden-Württemberg (Germany), two strains of the provisional taxon 'Candidatus Phytoplasma asteris' were identified within a carrot plot. For further analysis, strains M8 and M33 underwent shotgun sequencing, utilising single-molecule-real-time (SMRT) long-read sequencing and sequencing-by-synthesis (SBS) paired-end short-read sequencing techniques. Hybrid assemblies resulted in complete de novo assemblies of two genomes harboring circular chromosomes and two plasmids. Analyses, including average nucleotide identity and sequence comparisons of established marker genes, confirmed the phylogenetic divergence of 'Ca. P. asteris' and a different assignment of strains to the 16S rRNA subgroup I-A for M33 and I-B for M8. These groups exhibited unique features, encompassing virulence factors and genes, associated with the mobilome. In contrast, pan-genome analysis revealed a highly conserved gene set related to metabolism across these strains. This analysis of the Aster Yellows (AY) group reaffirms the perception of phytoplasmas as bacteria that have undergone extensive genome reduction during their co-evolution with the host and an increase of genome size by mobilome.

Prevalence of a 'Candidatus Phytoplasma solani'-Related Strain Designated as New 16SrXII-P Subgroup over 'Candidatus Arsenophonus phytopathogenicus' in Sugar Beet in Eastern Germany.

Two phloem-limited pathogens, 'Candidatus Arsenophonus phytopathogenicus' and 'Candidatus Phytoplasma solani', threaten sugar beet production in France, Switzerland, and Germany. Previous studies of these pathogens in Germany had focused on its western and southern regions, leaving a knowledge gap about eastern Germany. Despite their importance, this study is the first to investigate phytoplasmas in sugar beet in Saxony-Anhalt, Germany. A phytoplasma strain related to 'Ca. P. solani' is found predominant in Saxony-Anhalt, unlike in France, where 'Ca. P. solani' has a minor role compared with 'Ca. A. phytopathogenicus'. The phytoplasma strain infecting sugar beet in Saxony-Anhalt was classified into a new subgroup designated as 16SrXII-P. The multilocus sequence analysis (MLSA) of nonribosomal genes of the novel phytoplasma strain showed that it is significantly different from the reference and all previously reported 'Ca. P. solani' strains including the strain from western Germany. Analyses of sugar beet samples from previous years confirmed the presence of the 16SrXII-P strain in sugar beet as early as 2020 and also in Bavaria in southern Germany. Based on 16S rDNA analysis, 'Ca. A. phytopathogenicus' in Saxony-Anhalt is identical to strains in sugar beet in other parts of Germany and France, as well as to a strain in potato from Germany. The presence and prevalence of two phytoplasmas in sugar beet in Germany suggest that more attention should be directed toward understanding phytoplasma infection in sugar beet in this country.

Complete Genome of "Candidatus Phytoplasma rubi" RS, a Phytopathogenic Bacterium Associated with Rubus Stunt Disease.

The phytoplasma "Candidatus Phytoplasma rubi" is associated with Rubus stunt disease. The complete genome was determined by assembling Oxford Nanopore Technologies system-derived long reads, with short-read polishing with Illumina reads. The genome of strain RS, from Germany, is organized in one circular chromosome with a length of 762 kb.

Physiological and genomic characterisation of Luteimonas fraxinea sp. nov., a bacterial species associated with trees tolerant to ash dieback.

A group of isolates of the genus Luteimonas was characterised, which represented a specific component of the healthy core microbiome of Fraxinus excelsior in forest districts with a high infection rate of H. fraxineus, the causal agent of ash dieback. Based on phylogenomic and phenotypic analyses, a clear differentiation from related Luteimonas species was shown. Comparisons of the overall genome relatedness indices with the closest phylogenetic neighbours resulted in values below the recommended species cut-off levels. In addition, differences in several physiological and chemotaxonomic traits allowed a clear demarcation from the type strains of closely related species. Conclusively, the strain group was considered to represent a novel species in the genus Luteimonas, for which the name Luteimonas fraxinea sp. nov. is proposed, with strain D4P002T (=DSM 113273T = LMG 32455T) as the type strain. A functional analysis of the genome revealed features particularly associated with attachment, biofilm production and motility, indicating the ability of D4P002T to effectively colonise ash leaves. In nursery trials, ash seedlings inoculated with this strain showed suppression of the pathogen over a period of three years. This effect was accompanied by a significant shift in the bacterial microbiome of the plants. Altogether, the exclusive occurrence in the microbiome of tolerant ash trees, the genetic background and the results of the inoculation experiment suggest that strain D4P002T may suppress the penetration and spreading of H. fraxineus in or on ash leaves via colonisation resistance or trigger a priming effect of plant defences against the pathogen.

Revision of the 'Candidatus Phytoplasma' species description guidelines.

The genus 'Candidatus Phytoplasma' was proposed to accommodate cell wall-less bacteria that are molecularly and biochemically incompletely characterized, and colonize plant phloem and insect vector tissues. This provisional classification is highly relevant due to its application in epidemiological and ecological studies, mainly aimed at keeping the severe phytoplasma plant diseases under control worldwide. Given the increasing discovery of molecular diversity within the genus 'Ca. Phytoplasma', the proposed guidelines were revised and clarified to accommodate those 'Ca. Phytoplasma' species strains sharing >98.65 % sequence identity of their full or nearly full 16S rRNA gene sequences, obtained with at least twofold coverage of the sequence, compared with those of the reference strain of such species. Strains sharing <98.65 % sequence identity with the reference strain but >98.65 % with other strain(s) within the same 'Ca. Phytoplasma' species should be considered related strains to that 'Ca. Phytoplasma' species. The guidelines herein, keep the original published reference strains. However, to improve 'Ca. Phytoplasma' species assignment, complementary strains are suggested as an alternative to the reference strains. This will be implemented when only a partial 16S rRNA gene and/or a few other genes have been sequenced, or the strain is no longer available for further molecular characterization. Lists of 'Ca. Phytoplasma' species and alternative reference strains described are reported. For new 'Ca. Phytoplasma' species that will be assigned with identity ≥98.65 % of their 16S rRNA gene sequences, a threshold of 95 % genome-wide average nucleotide identity is suggested. When the whole genome sequences are unavailable, two among conserved housekeeping genes could be used. There are 49 officially published 'Candidatus Phytoplasma' species, including 'Ca. P. cocostanzaniae' and 'Ca. P. palmae' described in this manuscript.

The Genome Reduction Excludes the Ribosomal Rescue System in Acholeplasmataceae.

The trans-translation process is a ribosomal rescue system for stalled ribosomes processing truncated mRNA. The genes ssrA and smpB fulfil the key functions in most bacteria, but some species have either lost these genes or the function of the ribosomal rescue system is taken over by other genes. To date, the ribosomal rescue system has not been analysed in detail for the Acholeplasmataceae. This family, in the Mollicutes class, comprises the genus Acholeplasma and the provisional taxon "Candidatus Phytoplasma". Despite their monophyletic origin, the two clades can be separated by traits such as not representing primary pathogens for acholeplasmas versus being phytopathogenic for the majority of phytoplasmas. Both taxa share reduced genomes, but only phytoplasma genomes are characterised by a remarkable level of instability and reduction. Despite the general relevance of the ribosomal rescue system, information is lacking on coding, the genomic context and pseudogenisation of smpB and ssrA and their possible application as a phylogenetic marker. Herein, we provide a comprehensive analysis of the ribosomal rescue system in members of Acholeplasmataceae. The examined Acholeplasmataceae genomes encode a ribosomal rescue system, which depends on tmRNA encoded by ssrA acting in combination with its binding protein SmpB. Conserved gene synteny is evident for smpB, while ssrA shows a less conserved genomic context. Analysis of the tmRNA sequences highlights the variability of proteolysis tag sequences and short conserved sites at the 5'- and 3'-ends. Analyses of smpB provided no hints regarding the coding of pseudogenes, but they did suggest its application as a phylogenetic marker of Acholeplasmataceae - in accordance with 16S rDNA topology. Sequence variability of smpB provides sufficient information for species assignment and phylogenetic analysis.

Application of TaqMan Real-Time PCR for Detecting 'Candidatus Arsenophonus Phytopathogenicus' Infection in Sugar Beet.

The γ-proteobacterium 'Candidatus Arsenophonus phytopathogenicus' is assigned as the major pathogen of "Syndrome des basses richesses", a sugar beet disease characterised by a reduction in the sugar content of taproots and biomass yield. Despite the economic impact of this bacteriosis, diagnostics for this important pathogen currently rely on end-point PCR detection. Herein, we introduce a TaqMan qPCR for diagnostics of the agent targeting genes encoding a heat shock protein of the Hsp20 family and mannose-6-phosphate isomerase. Quantitation with synthetic oligonucleotides as standard showed that the developed TaqMan qPCR assays enable the detection of up to 100 target copies. A comparison between the TaqMan qPCR and end-point PCR for 'Ca. A. phytopathogenicus' detection was carried out on 78 sugar beet samples from different locations in southern Germany. The newly developed assays enable the fast, reliable and sensitive detection of 'Ca. A. phytopathogenicus' in sugar beet.

Multilocus Genotyping of 'Candidatus Phytoplasma solani' Associated with Rubbery Taproot Disease of Sugar Beet in the Pannonian Plain.

Rubbery taproot disease of sugar beet (RTD), associated with 'Candidatus Phytoplasma solani', appeared in 2020 on an epidemic scale in northern Serbia and southern Slovakia, situated at opposite edges of the Pannonian Plain. In the affected locations where the disease was assessed, symptomatic sugar beets were analysed for phytoplasma infection. Additionally, multilocus sequence analyses of 'Ca. P. solani' strains on epidemiologically informative marker genes (tuf, stamp and vmp1) were performed. Symptomatic sugar beets from other countries of the Pannonian Plain (Croatia, Hungary and Austria), one sample from Germany, and red beets from Serbia were included in the analyses. 'Ca. P. solani' was detected in sugar beet in all assessed countries, as well as in red beet. Molecular analyses revealed the high genetic variability of 'Ca. P. solani' with the presence of all four tuf-types (a, b1, b2 and d), 14 stamp genotypes (seven new) and five vmp1 profiles (one new). The most common multilocus genotype in Serbia, Slovakia, Croatia, and Hungary was dSTOLg (tuf-d/STOL/V2-TA). It was dominant on sites with epidemic RTD outbreaks in the Pannonian Plain and in several sugar beet fields with non-epidemic RTD occurrence suggesting the prevalence of a particular epidemiological pathway during the epidemic's phases.

Genomic Analysis of the Endophytic Stenotrophomonas Strain 169 Reveals Features Related to Plant-Growth Promotion and Stress Tolerance.

Plant-associated Stenotrophomonas isolates have great potential for plant growth promotion, especially under stress conditions, due to their ability to promote tolerance to abiotic stresses such as salinity or drought. The endophytic strain Stenotrophomonas sp. 169, isolated from a field-grown poplar, increased the growth of inoculated in vitro plants, with a particular effect on root development, and was able to stimulate the rooting of poplar cuttings in the greenhouse. The strain produced high amounts of the plant growth-stimulating hormone auxin under in vitro conditions. The comparison of the 16S rRNA gene sequences and the phylogenetic analysis of the core genomes showed a close relationship to Stenotrophomonas chelatiphaga and a clear separation from Stenotrophomonas maltophilia. Whole genome sequence analysis revealed functional genes potentially associated with attachment and plant colonization, growth promotion, and stress protection. In detail, an extensive set of genes for twitching motility, chemotaxis, flagella biosynthesis, and the ability to form biofilms, which are connected with host plant colonization, could be identified in the genome of strain 169. The production of indole-3-acetic acid and the presence of genes for auxin biosynthesis pathways and the spermidine pathway could explain the ability to promote plant growth. Furthermore, the genome contained genes encoding for features related to the production of different osmoprotective molecules and enzymes mediating the regulation of stress tolerance and the ability of bacteria to quickly adapt to changing environments. Overall, the results of physiological tests and genome analysis demonstrated the capability of endophytic strain 169 to promote plant growth. In contrast to related species, strain 169 can be considered non-pathogenic and suitable for biotechnology applications.

Rubbery Taproot Disease of Sugar Beet in Serbia Associated with 'Candidatus Phytoplasma solani'.

Rubbery taproot disease (RTD) of sugar beet was observed in Serbia for the first time in the 1960s. The disease was already described in neighboring Bulgaria and Romania at the time but it was associated with abiotic factors. In this study on RTD of sugar beet in its main growing area of Serbia, we provide evidence of the association between 'Candidatus Phytoplasma solani' (stolbur phytoplasma) infection and the occurrence of typical RTD symptomatology. 'Ca. P. solani' was identified by PCR and the sequence analyses of 16S ribosomal RNA, tuf, secY, and stamp genes. In contrast, the causative agent of the syndrome "basses richesses" of sugar beet-namely, 'Ca. Arsenophonus phytopathogenicus'-was not detected. Sequence analysis of the stolbur strain's tuf gene confirmed a previously reported and a new, distinct tuf stolbur genotype (named 'tuf d') that is prevalent in sugar beet. The sequence signatures of the tuf gene as well as the one of stamp both correlate with the epidemiological cycle and reservoir plant host. This study provides knowledge that, for the first time, enables the differentiation of stolbur strains associated with RTD of sugar beet from closely related strains, thereby providing necessary information for further epidemiological work seeking to identify insect vectors and reservoir plant hosts. The results of this study indicate that there are differences in hybrid susceptibility. Clarifying the etiology of RTD as a long-known and economically important disease is certainly the first step toward disease management in Serbia and neighboring countries.

Analyzing Ash Leaf-Colonizing Fungal Communities for Their Biological Control of Hymenoscyphus fraxineus.

The invasive ascomycete Hymenoscyphus fraxineus has been threatening Fraxinus excelsior populations throughout Europe for over two decades. Since the infection and first colonization by the pathogen occurs in leaves, leaf-colonizing microorganisms have been discussed as a barrier and as possible biocontrol agents against the disease. To identify fungal groups with health-supporting potential, we compared the fungal microbiota of compound leaves from susceptible and tolerant ash trees in four ash stands with high H. fraxineus exposure. The fungal communities were analyzed both culture-independently by ITS2 amplicon sequencing and by the taxonomic classification of 1,704 isolates using matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) or sequencing of the entire ITS region. The fungal community structure did not show significant differences depending on the health status. However, for several OTUs and a MALDI group, a significantly higher abundance was found in tolerant ash trees. Thus, the yeast Papiliotrema flavescens was significantly increased and accounted for 12.3% of the mycobiome of tolerant ashes (OTU0003), and it had also a distinctly higher abundance among the isolates. The filamentous ascomycete Sarocladium strictum was increased 24-fold among the isolates of tolerant trees, but its abundance was comparably low. An in vitro screening for the growth inhibition of the pathogen via cocultivation resulted in 28 yeast-like isolates and 79 filamentous fungi with antagonistic activity. A statistical cocultivation test on two H. fraxineus strains confirmed six of the yeast-like isolates that suppressed H. fraxineus significantly, from 39-50%, two of them through a fungicidal effect. The highest inhibition rates among the yeasts were found for three isolates belonging to Aureobasidium pullulans and P. flavescens. The cocultivation test of the filamentous isolates revealed higher effects compared to the yeasts. Four isolates showed significant inhibition of both H. fraxineus strains with a rate of 72-100%, and five further isolates inhibited only one H. fraxineus strain significantly. The most effective isolates were members of the genus Cladosporium. During the next step, in planta tests will be necessary to verify the efficacy of the antagonistic isolates and to assess their suitability as biocontrol agents.

Draft Genome Sequence of "Candidatus Phytoplasma pruni" (X-Disease Group, Subgroup 16SrIII-B) Strain ChTDIII from Argentina.

Herein, we report the draft genome sequence of "Candidatus Phytoplasma pruni" strain ChTDIII (subgroup 16SrIII-B). The final assembly consists of 790,517 nucleotides organized in 67 contigs (minimal size, 1 kb), with a G+C content of 29.4% and encoding 672 proteins.

A Comparative Analysis of Ash Leaf-Colonizing Bacterial Communities Identifies Putative Antagonists of Hymenoscyphus fraxineus.

In the last few years, the alarming spread of Hymenoscyphus fraxineus, the causal agent of ash dieback, has resulted in a substantial threat to native ash stands in central and northern Europe. Since leaves and leaf petioles are the primary infection sites, phyllosphere microorganisms are presumed to interact with the pathogen and are discussed as a source of biocontrol agents. We studied compound leaves from susceptible and visible infection-free trees in four ash stands with a high likelihood of infection to assess a possible variation in the bacterial microbiota, depending on the health status of the trees. The bacterial community was analyzed by culture-independent 16S rRNA gene amplicon sequencing and through the isolation and taxonomic classification of 2,589 isolates using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The bacterial community structure did not show significant differences. However, a set of amplicon sequence variants (ASVs) and MALDI groups belonging to Luteimonas, Aureimonas, Pseudomonas, Bacillus, and Paenibacillus were distinctly increased in tolerant trees, which may be associated with the ability of the tree to resist the pathogen. The most obvious differences were observed for Luteimonas, a genus that is also exclusively present in the healthy core microbiome. In a first in vitro screen of antagonists, approximately 11% of total isolates suppressed the growth of H. fraxineus, but a statistical test with two different H. fraxineus strains confirmed only the antagonistic activity of 8% of these isolates. The antagonistic isolates were assigned to Bacillus velezensis, Pantoea vagans, and Pseudomonas caspiana. Overall, our study provides a set of isolates or phylogenetic groups that might be involved in the process that prevents the penetration and spread of H. fraxineus. In the next step, in planta experiments are required with a longer period of exposure to H. fraxineus to evaluate effective isolates or consortia of isolates acting through direct antagonism or competition or indirectly by inducing resistance.

Widespread occurrence of 'Candidatus Phytoplasma ulmi' in elm species in Germany.

BACKGROUND: 'Candidatus Phytoplasma ulmi' is the agent associated with elm yellows and has been categorised in the European Union as a quarantine pathogen. For central and northern European countries, information on the occurrence and distribution of the pathogen and its impact on elms is scarce, so a survey of native elm trees has been conducted in Germany. RESULTS: About 6500 samples from Ulmus minor, Ulmus laevis and Ulmus glabra, were collected nationwide. Phytoplasma detection was performed by applying a universal 16Sr DNA-based quantitative PCR (qPCR) assay and a novel 'Ca. P. ulmi' specific qPCR assay targeting the 16S-23S spacer region. Both assays revealed that 28% of the samples were infected by 'Ca. P. ulmi', but infection rates of the elm species and regional incidences differed. The phytoplasma presence in the trees was not correlated to disease-specific symptoms. The survey identified a regional disparity of infection which was high in east, south and central Germany, whereas only a few infected sites were found in the western and northern parts of the country. Monitoring the seasonal titre of 'Ca. P. ulmi' in an infected tree by qPCR revealed a high colonisation in all parts of the tree throughout the year. CONCLUSIONS: 'Ca. P. ulmi' is widely present in elms in Germany. The rare occurrence of symptoms indicates either a high degree of tolerance in elm populations or a low virulence of pathogen strains enabling high infection rates in a long-living host.

Chamomile Floricolous Downy Mildew Caused by Peronospora radii.

Floricolous downy mildews (Peronospora, oomycetes) are a small, monophyletic group of mostly inconspicuous plant pathogens that induce symptoms exclusively on flowers. Characterization of this group of pathogens, and information about their biology, is particularly sparse. The recurrent presence of a disease causing flower malformation which, in turn, leads to high production losses of the medicinal herb Matricaria chamomilla in Serbia has enabled continuous experiments focusing on the pathogen and its biology. Peronospora radii was identified as the causal agent of the disease, and morphologically and molecularly characterized. Diseased chamomile flowers showed severe malformations of the disc and ray florets, including phyllody and secondary inflorescence formation, followed by the onset of downy mildew. Phylogeny, based on internal transcribed spacer and cox2, indicates clustering of the Serbian P. radii with other P. radii from chamomile although, in cox2 analyses, they formed a separate subcluster. Evidence pointing to systemic infection was provided through histological and molecular analyses, with related experiments validating the impact of soilborne and blossom infections. This study provides new findings in the biology of P. radii on chamomile, thus enabling the reconstruction of this floricolous Peronospora species' life cycle.

Genome and catabolic subproteomes of the marine, nutritionally versatile, sulfate-reducing bacterium Desulfococcus multivorans DSM 2059.

BACKGROUND: Sulfate-reducing bacteria (SRB) are key players of the carbon- and sulfur-cycles in the sediments of the world's oceans. Habitat relevant SRBs are often members of the Desulfosarcina-Desulfococcus clade belonging to the deltaproteobacterial family of Desulfobacteraceae. Despite this environmental recognition, their molecular (genome-based) physiology and their potential to contribute to organic carbon mineralization as well as to adapt to changing environmental conditions have been scarcely investigated. A metabolically versatile representative of this family is Desulfococcus multivorans that is able to completely oxidize (to CO2) a variety of organic acids, including fatty acids up to C14, as well as aromatic compounds. RESULTS: In this study the complete 4.46 Mbp and manually annotated genome of metabolically versatile Desulfococcus multivorans DSM 2059 is presented with particular emphasis on a proteomics-driven metabolic reconstruction. Proteomic profiling covered 17 substrate adaptation conditions (6 aromatic and 11 aliphatic compounds) and comprised 2D DIGE, shotgun proteomics and analysis of the membrane protein-enriched fractions. This comprehensive proteogenomic dataset allowed for reconstructing a metabolic network of degradation pathways and energy metabolism that consists of 170 proteins (154 detected; ~91 % coverage). Peripheral degradation routes feed via central benzoyl-CoA, (modified) ß-oxidation or methylmalonyl-CoA pathways into the Wood-Ljungdahl pathway for complete oxidation of acetyl-CoA to CO2. Dissimilatory sulfate reduction is fueled by a complex electron transfer network composed of cytoplasmic components (e.g., electron transfer flavoproteins) and diverse membrane redox complexes (Dsr, Qmo, Hmc, Tmc, Qrc, Nuo and Rnf). Overall, a high degree of substrate-specific formation of catabolic enzymes was observed, while most complexes involved in electron transfer appeared to be constitutively formed. CONCLUSIONS: A highly dynamic genome structure in combination with substrate-specifically formed catabolic subproteomes and a constitutive subproteome for energy metabolism and electron transfer appears to be a common trait of Desulfobacteraceae members.

Multi-omics analysis of niche specificity provides new insights into ecological adaptation in bacteria.

Different lifestyles, ranging from a saprophyte to a pathogen, have been reported in bacteria of one species. Here, we performed genome-wide survey of the ecological adaptation in four Burkholderia seminalis strains, distinguished by their origin as part of the saprophytic microbial community of soil or water but also including human and plant pathogens. The results indicated that each strain is separated from the others by increased fitness in medium simulating its original niche corresponding to the difference between strains in metabolic capacities. Furthermore, strain-specific metabolism and niche survival was generally linked with genomic variants and niche-dependent differential expression of the corresponding genes. In particular, the importance of iron, trehalose and d-arabitol utilization was highlighted by the involvement of DNA-methylation and horizontal gene transfer in niche-adapted regulation of the corresponding operons based on the integrated analysis of our multi-omics data. Overall, our results provided insights of niche-specific adaptation in bacteria.

Complete Genome Sequences of the Obligate Symbionts "Candidatus Sulcia muelleri" and "Ca. Nasuia deltocephalinicola" from the Pestiferous Leafhopper Macrosteles quadripunctulatus (Hemiptera: Cicadellidae).

Two bacterial symbionts of the European pest leafhopper, Macrosteles quadripunctulatus (Hemiptera: Cicadellidae), were fully sequenced. "Candidatus Sulcia muelleri" and "Ca. Nasuia deltocephalinicola" represent two of the smallest known bacterial genomes at 190 kb and 112 kb, respectively. Genome sequences are nearly identical to strains reported from the closely related host species, M. quadrilineatus.

'Candidatus Phytoplasma phoenicium' associated with almond witches'-broom disease: from draft genome to genetic diversity among strain populations.

BACKGROUND: Almond witches'-broom (AlmWB), a devastating disease of almond, peach and nectarine in Lebanon, is associated with 'Candidatus Phytoplasma phoenicium'. In the present study, we generated a draft genome sequence of 'Ca. P. phoenicium' strain SA213, representative of phytoplasma strain populations from different host plants, and determined the genetic diversity among phytoplasma strain populations by phylogenetic analyses of 16S rRNA, groEL, tufB and inmp gene sequences. RESULTS: Sequence-based typing and phylogenetic analysis of the gene inmp, coding an integral membrane protein, distinguished AlmWB-associated phytoplasma strains originating from diverse host plants, whereas their 16S rRNA, tufB and groEL genes shared 100 % sequence identity. Moreover, dN/dS analysis indicated positive selection acting on inmp gene. Additionally, the analysis of 'Ca. P. phoenicium' draft genome revealed the presence of integral membrane proteins and effector-like proteins and potential candidates for interaction with hosts. One of the integral membrane proteins was predicted as BI-1, an inhibitor of apoptosis-promoting Bax factor. Bioinformatics analyses revealed the presence of putative BI-1 in draft and complete genomes of other 'Ca. Phytoplasma' species. CONCLUSION: The genetic diversity within 'Ca. P. phoenicium' strain populations in Lebanon suggested that AlmWB disease could be associated with phytoplasma strains derived from the adaptation of an original strain to diverse hosts. Moreover, the identification of a putative inhibitor of apoptosis-promoting Bax factor (BI-1) in 'Ca. P. phoenicium' draft genome and within genomes of other 'Ca. Phytoplasma' species suggested its potential role as a phytoplasma fitness-increasing factor by modification of the host-defense response.