No abstract

The biocontrol effect of the non-pathogenic Fusarium oxysporum strain CS-20 against the tomato wilt pathogen F. oxysporum f. sp. lycopersici (FOL) has been previously reported to be primarily plant-mediated. This study shows that CS-20 produces proteins, which elicit defense responses in tomato plants. Three protein-containing fractions were isolated from CS-20 biomass using size exclusion chromatography. Exposure of seedling roots to one of these fractions prior to inoculation with pathogenic FOL strains significantly reduced wilt severity. This fraction initiated an ion exchange response in cultured tomato cells resulting in a reversible alteration of extracellular pH; increased tomato chitinase activity, and induced systemic resistance by enhancing PR-1 expression in tomato leaves. Two other protein fractions were inactive in seedling protection. The main polypeptide (designated CS20EP), which was specifically present in the defense-inducing fraction and was not detected in inactive protein fractions, was identified. The nucleotide sequence encoding this protein was determined, and its complete amino acid sequence was deduced from direct Edman degradation (25 N-terminal amino acid residues) and DNA sequencing. The CS20EP was found to be a small basic cysteine-rich protein with a pI of 9.87 and 23.43% of hydrophobic amino acid residues. BLAST search in the NCBI database showed that the protein is new; however, it displays 48% sequence similarity with a hypothetical protein FGSG_10784 from F. graminearum strain PH-1. The contribution of CS20EP to elicitation of tomato defense responses resulting in wilt mitigating is discussed.

No abstract

Much of the mitogenome variation observed in fungal lineages seems driven by mobile genetic elements (MGEs), which have invaded their genomes throughout evolution. The variation in the distribution and nucleotide diversity of these elements appears to be the main distinction between different fungal taxa, making them promising candidates for diagnostic purposes. Fungi of the genus Fusarium display a high variation in MGE content, from MGE-poor (Fusarium oxysporum and Fusarium fujikuroi species complex) to MGE-rich mitogenomes found in the important cereal pathogens F. culmorum and F. graminearum sensu stricto. In this study, we investigated the MGE variation in these latter two species by mitogenome analysis of geographically diverse strains. In addition, a smaller set of F. cerealis and F. pseudograminearum strains was included for comparison. Forty-seven introns harboring from 0 to 3 endonucleases (HEGs) were identified in the standard set of mitochondrial protein-coding genes. Most of them belonged to the group I intron family and harbored either LAGLIDADG or GIY-YIG HEGs. Among a total of 53 HEGs, 27 were shared by all fungal strains. Most of the optional HEGs were irregularly distributed among fungal strains/species indicating ancestral mosaicism in MGEs. However, among optional MGEs, one exhibited species-specific

Thymol, a secondary plant metabolite possessing antifungal and chemosensitizing activities, disrupts cell wall or membrane integrity and interferes with ergosterol biosynthesis. Thymol also functions as a redox-active compound inducing generation of reactive oxygen species and lipid peroxidation in fungal cells. Previously, we showed thymol significantly enhanced the in vitro growth inhibitory effect of difenoconazole against Bipolaris sorokiniana and Parastagonospora nodorum. More recently, we demonstrated a possibility to use thymol to overcome the resistance of a P. nodorum strain able to grow on difenoconazole-containing media. However, potential for thymol to serve as a chemosensitizing agent in seed or plant treatments, to provide an effective suppression of the above-mentioned plant pathogens by triazole fungicides applied in lowered dosages, had yet to be tested. In the work presented here, we showed combined treatments of naturally infected barley seeds with thymol and difenoconazole (Dividend® 030 FS) synergistically exacerbated the protective effect against common root rot agent, B. sorokiniana, and other fungi (Fusarium spp. and Alternaria spp.). Similarly, co-applied treatment of wheat seeds, artificially inoculated with Fusarium culmorum, resulted in equivalent reduction of disease incidence on barley seedlings as application of Dividend®, alone, at a ten-fold higher dosage. In foliar treatments of wheat seedlings, thymol combined with Folicur® 250 EC (a.i. tebuconazole) enhanced sensitivity of P. nodorum, a glume/leaf blotch pathogen, to the fungicide and provided a significant mitigation of disease severity on treated seedlings, compared to controls, without increasing Folicur® dosages. Folicur® co-applied with thymol was also significantly more effective against a strain of P. nodorum tolerant to Folicur® alone. No additional deoxynivalenol or zearalenone production was found when a toxigenic F. culmorum was cultured in a nutrient medium containing thymol at a concentration used for chemosensitization of root rot agents. Accordingly, F. culmorum exposure to thymol at the sensitizing concentration did not up-regulate key genes associated with the biosynthesis of trichothecene or polyketide mycotoxins in this pathogen. Further studies using field trials are necessary to determine if thymol-triazole co-applications result in sensitization of seed- and foliar-associated plant pathogenic fungi, and if thymol affects production of fusarial toxins under field conditions.

The recent taxonomic diversification of bacterial genera Pectobacterium and Dickeya, which cause soft rot in plants, focuses attention on the need for improvement of existing methods for the detection and differentiation of these phytopathogens. This research presents a whole genome-based approach to the selection of marker sequences unique to particular species of Pectobacterium. The quantitative real-time PCR assay developed is selective in the context of all tested Pectobacterium atrosepticum strains and is able to detect fewer than 102 copies of target DNA per reaction. The presence of plant DNA extract did not affect the sensitivity of the assay.

We performed a three-locus phylogenetic analysis of Fusarium strains presumably capable of trichothecene production, which were deposited in the Russian national collections. The intra- and interspecific polymorphism of partial sequences of the translation elongation factor 1 alpha (TEF1α) gene and two genes from the trichothecene cluster TRI5 and TRI14 was studied. A study of 60 strains of different origins using DNA markers confirmed, and in the case for several strains, clarified their taxonomic characteristics. As a result, a strain of F. commune (F-900) was identified in Russia for the first time. Furthermore, the strain F-846 proved to be phylogenetically distinct from any of the known Fusarium species. F. equiseti strains from Northwest Russia were found to belong to the North European group (I), whereas a strain from the North Caucasus – to the South European one (II). Partial TRI14 sequences from 9 out of 12 species were determined for the first time. Their comparative analysis demonstrated a relatively high level of intraspecific variability in F. graminearum and F. sporotrichioides, but no correlation between the sequence polymorphism and the geographic origin of the strains or their chemotype was found. Specific chemotypes of trichothecene B producers were characterized using two primer sets. The chemotyping results were verified by HPLC.

Fungal complexes are often composed of morphologically nearly indistinguishable species with high genetic similarity. However, despite their close relationship, they can exhibit distinct phenotypic differences in pathogenicity and production of mycotoxins. Many plant pathogenic and toxigenic fungi have been shown to consist of such cryptic species. Identification of cryptic species in economically important pathogens has added value in epidemiologic studies and provides opportunities for better control. Analysis of mitochondrial genomes or mitogenomics opens up dimensions for improved diagnostics of fungi, especially when efficient recovery of DNA is problematic. In comparison to nuclear DNA, mitochondrial DNA (mtDNA) can be amplified with improved efficacy due to its multi-copy nature. However, to date, only a few studies have demonstrated the usefulness of mtDNA for identification of cryptic species within fungal complexes. In this study, we explored the value of mtDNA for identification of one of the most important cereal pathogens Fusarium graminearum sensu stricto (F.g.). We found that homing endonucleases (HEGs), which are widely distributed in mitogenomes of fungi, display small indel polymorphism, proven to be potentially species specific. The resulting small differences in their lengths may facilitate further differentiation of F.g. from the other cryptic species belonging to F. graminearum species complex. We also explored the value of SNP analysis of the mitogenome for typing F.g. The success in identifying F.g. strains was estimated at 96%, making this tool an attractive complement to other techniques for identification of F.g.