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Povzetek: Phytoplasmas of the 16SrIII group are wide spread, and have a broad plant host range. Among these, ‘Candidatus phytoplasma pruni’ (‘Ca. P. pruni’; phytoplasmas of 16SrIII subgroup A) can cause serious diseases in Prunus species and ‘Ca. P. pruni’-related strains can infect other plant species, including grapevines. In this study, a new real-time PCR detection system was developed for ‘Ca. P. pruni’ using TaqMan chemistry. This test was designed to detect ‘Ca. P. pruni’, by amplifying the species-specific secY gene. In addition, a test to amplify the group-specific 16S rRNA gene region was also developed. The performances of both tests were evaluated. The test that amplifies the secY gene provided reliable and quick detection of ‘Ca. P. pruni’. Using the newly developed and validated test, ‘Ca. P. pruni’ was not found in any of the 434 field samples collected from different plants species grown in different regions of Slovenia.
Ključne besede: phytoplasma, X-disease, real-time PCR, Prunus
Objavljeno v DiRROS: 22.07.2024; Ogledov: 139; Prenosov: 141
Celotno besedilo (2,00 MB)
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Povzetek: High-throughput sequencing (HTS), more specifically RNA sequencing of plant tissues, has become an indispensable tool for plant virologists to detect and identify plant viruses. During the data analysis step, plant virologists typically compare the obtained sequences to reference virus databases. In this way, they are neglecting sequences without homologies to viruses, which usually represent the majority of sequencing reads. We hypothesized that traces of other pathogens might be detected in this unused sequence data. In the present study, our goal was to investigate whether total RNA-seq data, as generated for plant virus detection, is also suitable for the detection of other plant pathogens and pests. As proof of concept, we first analyzed RNA-seq datasets of plant materials with confirmed infections by cellular pathogens in order to check whether these non-viral pathogens could be easily detected in the data. Next, we set up a community effort to re-analyze existing Illumina RNA-seq datasets used for virus detection to check for the potential presence of non-viral pathogens or pests. In total, 101 datasets from 15 participants derived from 51 different plant species were re-analyzed, of which 37 were selected for subsequent in-depth analyses. In 29 of the 37 selected samples (78%), we found convincing traces of non-viral plant pathogens or pests. The organisms most frequently detected in this way were fungi (15/37 datasets), followed by insects (13/37) and mites (9/37). The presence of some of the detected pathogens was confirmed by independent (q)PCRs analyses. After communicating the results, 6 out of the 15 participants indicated that they were unaware of the possible presence of these pathogens in their sample(s). All participants indicated that they would broaden the scope of their bioinformatic analyses in future studies and thus check for the presence of non-viral pathogens. In conclusion, we show that it is possible to detect non-viral pathogens or pests from total RNA-seq datasets, in this case primarily fungi, insects, and mites. With this study, we hope to raise awareness among plant virologists that their data might be useful for fellow plant pathologists in other disciplines (mycology, entomology, bacteriology) as well.
Ključne besede: plant viruses, plant virus detection, plant virology, high-throughput sequencing, RNA sequencing, plant tissues, plant pathogen, diagnostics, high-throughput sequencing, metagenomics, metatranscriptomics
Objavljeno v DiRROS: 12.07.2024; Ogledov: 142; Prenosov: 114
Celotno besedilo (1,70 MB)

Povzetek: Flavescence dorée (FD) phytoplasma from 16SrV-C and -D subgroups cause severe damage to grapevines throughout Europe. This phytoplasma is transmitted from grapevine to grapevine by the sap-sucking leafhopper Scaphoideus titanus. European black alder and clematis serve as perennial plant reservoirs for 16SrV-C phytoplasma strains, and their host range has recently been extended to hazelnuts. In Slovenia, hazelnut orchards are declining due to 16SrV phytoplasma infections, where large populations of the non-autochthonous leafhopper Orientus ishidae have been observed. To better characterise the phytoplasma-induced decline of hazelnut and possible transmission fluxes between these orchards and grapevine, genetic diversity of 16SrV phytoplasmas in grapevine, hazelnut and leafhoppers was monitored from 2017 to 2022. The nucleotide sequence analysis was based on the map gene. The most prevalent map genotype in grapevine in all wine-growing regions of Slovenia was M54, which accounted for 84 % of the 176 grapevines tested. Besides M54, other epidemic genotypes with lower frequency were M38 (6 %), M51 (3 %), M50 (2 %) and M122 (1 %). M38, M50 and M122 were also detected in infected cultivated hazelnuts and in specimens of O. ishidae leafhopper caught in declining hazelnut orchards. It suggests that this polyphagous vector could be responsible for phytoplasma infection in hazelnut orchards and possibly for some phytoplasma exchanges between hazelnuts and grapevine. We hereby describe new genotypes: M158 in grapevine as well as four never reported genotypes M159 to M162 in hazelnut. Of these four genotypes in hazelnut, one (M160) was also detected in O. ishidae. Analysis of additional genes of the new genotypes allowed us to assign them to the VmpA-III cluster, which corresponds to the 16SrV-C strains previously shown to be compatible with S. titanus transmission.
Ključne besede: phytoplasmas, haselnuts, grapevine, Flavescence dorée, epidemiology, plant disease
Objavljeno v DiRROS: 26.07.2023; Ogledov: 645; Prenosov: 470
Celotno besedilo (3,82 MB)
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