Kube, Michael


Publications (18)

Genome Comparison of ‘Candidatus Phytoplasma rubi’ with Genomes of Other 16SrV Phytoplasmas Highlights Special Group Features

Citation
Böhm et al. (2023). Applied Microbiology 3 (3)
Names (2)
Ca. Phytoplasma ziziphi Ca. Phytoplasma rubi
Subjects
Industrial and Manufacturing Engineering Mechanical Engineering Metals and Alloys Strategy and Management
Abstract
Phytoplasmas are associated with important bacterial diseases, causing severe symptoms in agricultural and ornamental crops. ‘Candidatus Phytoplasma rubi’, associated with the Rubus stunt in raspberries (Rubus idaeus) and blackberries (Rubus subgenus Rubus), causes shortened internodes, elongated sepals, proliferation, phyllody, and virescence. The recently published genome of ‘Ca. P. rubi’ RS enabled a comprehensive genomic comparison to the complete genomes of 16SrV phytoplasmas, comprising strains of the flavescence dorée-associated phytoplasma CH and two ‘Candidatus Phytoplasma ziziphi’ strains. Besides the typical transporters and metabolic features of phytoplasmas, the phosphorolysis of sucrose and the utilization of the carboxylic acid L-lactate became apparent for the 16SrV-group. With respect to the effector repertoire and the encoded immunodominant membrane proteins involved in host colonization, the group revealed conserved features that comprise the variable membrane proteins A and B. However, SAP11- and SAP54 orthologs were limited to ‘Ca. P. rubi’ RS and ‘Ca. P. ziziphi’. Genome-sequence-based phylogenetic analysis supports the close relationship of these genomes relative to alder yellows phytoplasmas. The analyses supported the impact of the mobilome on phytoplasma evolution but also highlighted that there is the possibility of identifying phytoplasmas with a larger metabolic repertoire in the future.

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

Citation
Duckeck et al. (2023). Microbiology Resource Announcements 12 (5)
Names (1)
Ca. Phytoplasma rubi
Subjects
Genetics Immunology and Microbiology (miscellaneous) Molecular Biology
Abstract
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.

Prevalence of a 'Candidatus Phytoplasma solani'-related strain designated as new 16SrXII-P subgroup over 'Candidatus Arsenophonus phytopathogenicus' in sugar beet in eastern Germany

Citation
Duduk et al. (2023). Plant Disease
Names (2)
Ca. Phytoplasma solani Ca. Arsenophonus phytopathogenicus
Subjects
Agronomy and Crop Science Plant Science
Abstract
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 to 'Ca. A. phytopathogenicus'. The phytoplasma strain infecting sugar beet in Saxony-Anhalt was classified into a new subgroup designated as 16SrXII-P. The MLSA of non-ribosomal genes of the novel phytoplasma strain showed that it is significantly different from the reference and all previously reported 'Ca. P. solani' strains including 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 towards understanding phytoplasma infection in sugar beet in this country.

Revision of the ‘Candidatus Phytoplasma’ species description guidelines

Citation
Bertaccini et al. (2022). International Journal of Systematic and Evolutionary Microbiology 72 (4)
Names (1)
Ca. Phytoplasma
Subjects
Ecology, Evolution, Behavior and Systematics General Medicine Microbiology
Abstract
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.

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

Citation
Zübert, Kube (2021). Pathogens 10 (11)
Names (2)
Ca. Arsenophonus phytopathogenicus Arsenophonus
Subjects
General Immunology and Microbiology Immunology and Allergy Infectious Diseases Microbiology (medical) Molecular Biology
Abstract
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

Citation
Ćurčić et al. (2021). Microorganisms 9 (9)
Names (1)
Ca. Phytoplasma solani
Subjects
Microbiology Microbiology (medical) Virology
Abstract
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.

Rubbery Taproot Disease of Sugar Beet in Serbia Associated with ‘Candidatus Phytoplasma solani’

Citation
Ćurčić et al. (2021). Plant Disease 105 (2)
Names (1)
Ca. Phytoplasma solani
Subjects
Agronomy and Crop Science Plant Science
Abstract
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.

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

Citation
Fernández et al. (2020). Microbiology Resource Announcements 9 (38)
Names (1)
Ca. Phytoplasma pruni
Subjects
Genetics Immunology and Microbiology (miscellaneous) Molecular Biology
Abstract
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.

Widespread occurrence of ‘Candidatus Phytoplasma ulmi’ in elm species in Germany

Citation
Schneider et al. (2020). BMC Microbiology 20 (1)
Names (1)
Ca. Phytoplasma ulmi
Subjects
Microbiology Microbiology (medical)
Abstract
Abstract 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.