Fungal trunk diseases (FTDs) have been a significant threat to the global stone fruit industry. FTDs are caused by a consortium of wood-decaying fungi. These fungi colonize woody tissues, causing cankers, dieback, and other decline-related symptoms in host plants. In this study, a detailed screening of the fungal microbiota associated with the decline of stone fruit trees in the Czech Republic was performed. The wood fragments of plum and apricot trees showing symptoms of FTDs were subjected to fungal isolation. The partial internal transcribed spacer region, partial beta-tubulin, and translation elongation factor 1-α genes were amplified from genomic DNA extracted from fungal cultures. All isolates were classified, and the taxonomic placement of pathogenic strains was illustrated in phylogenetic trees. The most abundant pathogenic genus was Dactylonectria (31%), followed by Biscogniauxia (13%), Thelonectria (10%), Eutypa (9%), Dothiorella (7%), Diplodia (6%), and Diaporthe (6%). The most frequent endophytic genus was Aposphaeria (17%). The pathogenicity of six fungal species (Cadophora daguensis, Collophorina africana, Cytospora sorbicola, Dothiorella sarmentorum, Eutypa lata, and E. petrakii var. petrakii) to four Prunus spp. was evaluated, and Koch's postulates were fulfilled. All tested isolates caused lesions on at least one Prunus sp. The most aggressive species was E. lata, which caused the largest lesions on all four tested Prunus spp., followed by E. petrakii var. petrakii and D. sarmentorum. Japanese plum (Prunus salicina) and almond (P. amygdalus) were the most susceptible hosts, while apricot (P. armeniaca) was the least susceptible host in the pathogenicity trial.
- Klíčová slova
- FTDs, Prunus, apricot, fungi, molecular identification, pathogenicity, plum, stone fruit, taxonomy,
- MeSH
- DNA fungální genetika MeSH
- dřevo mikrobiologie MeSH
- elongační faktor 1 genetika MeSH
- fylogeneze * MeSH
- houby genetika klasifikace izolace a purifikace patogenita MeSH
- meruňka obecná * mikrobiologie genetika MeSH
- nemoci rostlin * mikrobiologie MeSH
- slivoň mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
- Názvy látek
- DNA fungální MeSH
- elongační faktor 1 MeSH
A long-term exposure of bacteria to zinc oxide and zinc oxide nanoparticles leads to major alterations in bacterial morphology and physiology. These included biochemical and physiological processes promoting the emergence of strains with multi-drug resistance and virulence traits. After the removal of zinc pressure, bacterial phenotype reversed back to the original state; however, certain changes at the genomic, transcriptomic, and proteomic level remained. Why is this important? The extensive and intensive use of supplements in animal feed effects the intestinal microbiota of livestock and this may negatively impact the health of animals and people. Therefore, it is crucial to understand and monitor the impact of feed supplements on intestinal microorganisms in order to adequately assess and prevent potential health risks.
- Klíčová slova
- antimicrobial resistance, genome, nanoparticles, phenotype, proteome, transcriptome, virulence, zinc, zinc oxide,
- MeSH
- Escherichia coli genetika MeSH
- lidé MeSH
- multiomika MeSH
- oxid zinečnatý * chemie MeSH
- proteomika MeSH
- zinek * farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- oxid zinečnatý * MeSH
- zinek * MeSH
The newly discovered Xanthomonas phage M29 (Xp M29) is the first lytic phage infecting Xanthomonas campestris pv. campestris (Xcc) that was isolated from cabbage leaves in the Czech Republic. The phage consists of icosahedral head approximately 60 nm in diameter and a probably contractile tail of 170 nm. The complete genome size was 42 891 bp, with a G + C content of 59.6%, and 69 ORFs were predicted on both strands. Pairwise nucleotide comparison showed the highest similarity with the recently described Xanthomonas phage FoX3 (91.2%). Bacteriophage Xp M29 has a narrow host range infecting 5 out of 21 isolates of Xcc. Xp M29 is a novel species in a newly formed genus Foxunavirus assigned directly to the class Caudoviricetes.
- Klíčová slova
- Bacteriophage, Biocontrol, Xanthomonas campestris pv. campestris whole-genome sequence,
- MeSH
- bakteriofágy * genetika MeSH
- Myoviridae MeSH
- Xanthomonas campestris * genetika MeSH
- Xanthomonas * genetika MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
During a previous study on microfungi associated with clematis roots, Penicillium-like fungi were isolated and identified based on morphology. In this study, we subjected those strains to a detailed examination which led to the proposal of two taxonomic novelties, named Rasamsonia chlamydospora and Talaromyces clematidis. The first taxon is characterized by rough-walled mycelium, acerose to flask shaped phialides, cylindrical conidia and by production of chlamydospore-like structures. The four-loci-based phylogeny analysis delineated the taxon as a taxonomic novelty in Rasamsonia. Talaromyces clematidis is characterized by restricted growth on Czapek yeast extract agar, dichloran 18% glycerol agar and yeast extract sucrose agar, and production of yellow ascomata on oatmeal agar. Phylogenetic analyses placed this taxon as a taxonomic novelty in Talaromyces sect. Bacillispori. Both taxa are introduced here with detailed descriptions, photoplates and information on their phylogenetic relationship with related species.
- MeSH
- agar MeSH
- Eurotiales * MeSH
- fylogeneze MeSH
- Talaromyces * genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
- Názvy látek
- agar MeSH
Recent developments in high-throughput sequencing (HTS) technologies and bioinformatics have drastically changed research in virology, especially for virus discovery. Indeed, proper monitoring of the viral population requires information on the different isolates circulating in the studied area. For this purpose, HTS has greatly facilitated the sequencing of new genomes of detected viruses and their comparison. However, bioinformatics analyses allowing reconstruction of genome sequences and detection of single nucleotide polymorphisms (SNPs) can potentially create bias and has not been widely addressed so far. Therefore, more knowledge is required on the limitations of predicting SNPs based on HTS-generated sequence samples. To address this issue, we compared the ability of 14 plant virology laboratories, each employing a different bioinformatics pipeline, to detect 21 variants of pepino mosaic virus (PepMV) in three samples through large-scale performance testing (PT) using three artificially designed datasets. To evaluate the impact of bioinformatics analyses, they were divided into three key steps: reads pre-processing, virus-isolate identification, and variant calling. Each step was evaluated independently through an original, PT design including discussion and validation between participants at each step. Overall, this work underlines key parameters influencing SNPs detection and proposes recommendations for reliable variant calling for plant viruses. The identification of the closest reference, mapping parameters and manual validation of the detection were recognized as the most impactful analysis steps for the success of the SNPs detections. Strategies to improve the prediction of SNPs are also discussed.
- Klíčová slova
- Bioinformatic, Genomic, Plant, Variant, Virus,
- MeSH
- genom virový genetika MeSH
- jednonukleotidový polymorfismus * genetika MeSH
- lidé MeSH
- výpočetní biologie MeSH
- vysoce účinné nukleotidové sekvenování * MeSH
- znalosti MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Viticulture is a traditional branch of agriculture in the Czech Republic. Grapevines (Vitis vinifera L.) are cultivated on more than 18,000 hectares in the wine-growing regions of Bohemia and South Moravia. South Moravia alone accounts for more than 90 % of the total wine-growing area in the country. Grapevine yellows are a complex of diseases associated with the phytoplasma presence. Phytoplasmas of at least five different groups can cause similar symptoms in grapevines, and they can be distinguished only on a molecular basis (EPPO 2016). One of them, the grapevine Flavescence dorée phytoplasma (GFDP), which belongs to the 16SrV group, is listed in Annex II, Part B, of the Commission Implementing Regulation (EU) 2019/2072 of 28 November 2019 as a Union quarantine pest known to occur in the Union territory. Official surveys for GFDP in the Czech Republic have been carried out since 2007. In 2016, the first occurrence of Scaphoideus titanus Ball, 1932, the main vector of GFDP, was reported in the South Moravian Region (EPPO Reporting Service 2016). This is a matter of concern because it indicates that there is a risk of disease dissemination to other geographical locations. In September 2021, a total of 250 samples of V. vinifera (preferentially focused on symptomatic plants) and four samples of the wild plant host Clematis vitalba L. were collected from 50 vineyards in South Moravia. Total DNA was extracted using High Pure PCR Template Preparation Kit (Roche, Basel, Switzerland). For phytoplasma screening, a real-time PCR test for generic detection of phytoplasmas was used (Christensen et al. 2004). Samples evaluated as positive were further tested by PCR using phytoplasma universal P1 and P7 primers (Deng and Hiruki 1991; Schneider et al. 1995), followed by nested PCR using the 16SrV group-specific primers fB1 and rULWS1 (Smart et al. 1996). For identification of 16SrV phytoplasma, sequence analysis of the secY-map genetic locus was performed. Two sets of primers were used: FD9f5/MAPr1 primers for the first PCR and FD9f6/MAPr2 for the nested PCR (Arnaud et al. 2007) with PCRBIO TaqMix (PCR Biosystems, London, UK). The nested PCR products were purified and sequenced (Eurofins Genomics, Ebersberg, Germany). The sequences were compared with sequences from the GenBank database. Phytoplasma of the 16SrV group was detected in three samples: V. vinifera cv. Gewürztraminer with symptoms of leaf reddening with no rolling and no other typical symptoms; C. vitalba L. with leaf curling (Fig. 1A); symptomless C. vitalba. The obtained sequences of the secY-map locus of all three 16SrV-positive samples were identical to the sequence of GFDP, isolate Vv-SI257 (Acc. No. FN811141), detected in grapevine in Tuscany (Italy), which belongs to 16SrV group. The sequence of the V. vinifera cv. Gewürztraminer isolate was submitted to GenBank under Acc. No. OQ185203. This isolate belongs to the Map-FD3 cluster (Fig. 1B), and the genotype identified is M51 (corresponding to FD-C), which has already been found in C. vitalba and outbreaks of Flavescence dorée in grapevines in some other European countries (Malembic-Maher et al. 2020). Based on the abovementioned results, this is the first report of the GFDP in the Czech Republic.
- Klíčová slova
- Clematis, GFDP, grapevine,
- Publikační typ
- časopisecké články MeSH
Neofusicoccum parvum (Pennycook & Samuels) Crous, Slippers & A.J.L. Phillips is a cosmopolitan pathogen causing dieback of multiple diverse woody hosts including highbush blueberry (Vaccinium corymbosum L.). This fungus can survive inside colonized plants without causing any symptoms for several years. Once the endophytic lifestyle is switched to the parasitic one, the symptoms of dieback can rapidly occur (bronze leaves, necroses under the bark, apoplexy) and the plant usually declines within a few weeks (Slipper and Wingfield 2007). In August 2022, blueberry plants displaying symptoms described above were observed in a production orchard located in Hovorany, the Czech Republic. Around 3 % of 1000 observed plants were symptomatic. In order to identify the pathogen, leaves, stems and roots of three diseased plants were collected, sectioned into small pieces (5 × 5 mm), surface sterilized (60 s in 75% ethanol, followed by 60 s in 1% sodium hypochlorite and rinsed three times using sterile distilled water), plated on potato dextrose agar (PDA) supplemented with 0.5 g/liter of streptomycin sulfate (PDAS) (Biosynth, Staad, Switzerland) and incubated at 25°C for 2 weeks at dark. Newly developed mycelia were immediately transferred to fresh PDA plates and purified by single-spore or hyphal-tip isolation. In total 33 fungal isolates were obtained. All the 33 isolates shared similar morphology and resembled Botryosphaeriaceae spp. Colonies on PDA (7 d at 25°C) were felty, white to iron grey in the centre. Conidiomata were observed on sterile pine needles on 2 % water agar (WA) at 25°C under near-UV light after 2 wks (110-220 × 60-175 μm). Conidia (n=30) were cylindrical to ellipsoidal, hyaline, 0(-1)-septate, (3.8-8.1 × 2-3 μm). Two representative isolates were deposited at the Westerdijk Fungal Biodiversity Institute, Utrecht, the Netherlands (CBS 149846 and CBS 149847). The partial internal transcribed spacer (ITS) regions, beta-tubulin gene (tub2) and translation elongation factor 1-alpha (tef) gene were amplified from genomic DNA of both isolates following primers and protocols previously described (Eichmeier et al. 2020). Newly generated sequences were deposited in NCBI GenBank (acc. nos. ITS: OQ376566, OQ376567; tub2: OQ401701, OQ401702 and tef: OQ401699, OQ401700), being >99% identical (ITS 483/484 nt, tub2 426/430 nt and tef 230/232 nt) with the ex-type ITS (AY236943), tub2 (AY236888) and tef (AY236917) sequences of N. parvum strain CMW9081. Phylogenetically, newly obtained isolates grouped with ex-type and another three cultures of N. parvum in the three gene-based phylogenetic tree with strong 98/1.0 (BP/PP) support. To confirm pathogenicity, one-year-old canes of ten two-year-old V. corymbosum plants grown in pots were wounded by a 5 mm diam cork borer, and a 5-mm mycelial plug of a 7-day-old culture of both (CBS 149846 and CBS 149847) strains (five plants per strain) was inserted into the wound. Ten plants were inoculated with sterile PDA plugs and served as controls. Wounds were covered by sterile wet cotton, sealed with Parafilm® and inoculated plants were maintained in a growth chamber at 20 °C with 12/12 h light/dark period. Within two weeks, inoculated shoots changed colour from green to dark brown and exhibited dark necroses under the bark; after one month inoculated plants declined, while controls remained symptomless. The pathogen was reisolated from the inoculated plants with 100 % re-isolation rate, and its identity confirmed by sequencing ITS region. The experiment was repeated. Neofusicoccum parvum causing dieback of highbush blueberry was already reported from Australia, California, Chile, China, Italy, Mexico, Portugal and Uruguay (Rossman et al. 2023). Pecenka et al. (2021) reported a presence of another pathogen - Lasiodiplodia theobromae (Pat.) Griffon & Maubl. from the same plantation. This suggests that stem blight and dieback of highbush blueberry is caused by more than one Botryosphaeriaceae spp. as it was previously proved by Xu et al. (2015). To the best of our knowledge, this is the first report of stem blight and dieback of highbush blueberry caused by N. parvum in the Czech Republic.
- Klíčová slova
- Botryosphaeriaceae, Causal Agent, Fungi, Neofusicoccum parvum, Vaccinium corymbosum, highbush blueberry, molecular identification, pathogenicity,
- Publikační typ
- časopisecké články MeSH
- Klíčová slova
- Candidatus Liberibacter asiaticus, Huanglongbing (HLB), Xanthomonas campestris, black rot disease, citrus, plant vascular system,
- Publikační typ
- úvodníky MeSH
Clematis L. is one of the largest genera of Ranunculaceae, accommodating over 300 plant species (Wang & Li 2005). They are mostly flowering creepers commonly grown as ornamentals. Clematis leaf spot and wilt is a fungal disease caused by Calophoma clematidina (Thüm.) Q. Chen & L. Cai. Infected plants initially show irregular brown to black leaf spots which later turn into large necroses, usually leading to wilt disease. In June 2021, Clematis plants displaying symptoms described above were observed in three independent nurseries located in three counties (Brno-venkov, Břeclav and Nymburk) in the Czech Republic. Around 60% of 120 inspected plants were symptomatic, including both mother plants and young plants. Leaves, stems and roots of 43 diseased plants originating from the three nurseries were collected, sectioned into small pieces (5 × 5 mm), surface sterilized (60 sec in 75% ethanol, followed by 60 sec in 1% sodium hypochlorite and rinsed three times using sterile distilled water), plated on potato dextrose agar (PDA) and incubated at 25°C for 5 weeks. Newly developed mycelia were immediately transferred to a fresh PDA plates and purified by single-spore isolation. A total of 21 strains morphologically resembled the genus Calophoma. Colonies on PDA (7 d at 25°C) were felty, white to olivaceous/iron grey in the centre. Conidiomata were dark brown, pycnidial, solitary or in groups, (117-220 × 65-170 μm). Conidia were cylindrical to ellipsoidal, hyaline, 0(-1)-septate, (4-8 × 2-3 μm). Two representative isolates were deposited at the Westerdijk Fungal Biodiversity Institute, Utrecht, the Netherlands (CBS 149230 and CBS 149231). The partial internal transcribed spacer (ITS) regions, large ribosomal subunit of the nrRNA gene (LSU), beta-tubulin gene (tub2) and RNA polymerase II second largest subunit gene (rpb2) were amplified from genomic DNA of both isolates following protocols previously described (Spetik et al. 2022). Sequences were deposited in NCBI GenBank (accession nos. ITS: ON107539, ON107540; LSU: ON108575, ON108576; tub2: ON314832, ON314833; rpb2: ON125007, ON125008), being 100% identical with that of the ex-type strain of C. clematidina (CBS 108.79), ITS (NR_135964), LSU (FJ515632), tub2 (FJ427100), and rpb2 (KT389588). Phylogenetically, the two representative isolates formed a fully supported clade with sequences of the ex-type and another culture of C. clematidina in the multigene phylogeny. To confirm Koch's postulates, leaves of ten two-month-old Clematis plants grown in pots were wounded by a needle and inoculated with a conidial suspension (1.0 × 106 conidia ml-1) of both strains (five plants per strain) following Golazar et al. (2011). Ten plants were mock-inoculated with sterile distilled water and served as controls. Within one month, inoculated plants exhibited dark necrotic leaf spots similar to the symptoms observed in the nurseries, while controls remained symptomless. Calophoma clematidina was reisolated from the inoculated plants, and its identity confirmed (ITS, GenBank OP363927). The experiment was repeated. Although known from Europe, this is the first report of Clematis leaf spot and wilt caused by C. clematidina in the Czech Republic. Clematis leaf spot and wilt represents a serious disease in Czech nurseries, with the pathogen present in leaves, stems and roots of Clematis spp.
- Klíčová slova
- Calophoma clematidina, Causal Agent, Clematis, Czech Republic, Fungi, molecular identification, nurseries,
- Publikační typ
- časopisecké články MeSH
