First Report of Globisporangium (Pythium) mastophorum causing Damping-off / Root Rot on Parsley in Slovenia.

Pythium-like species cause damping-off symptoms of various hosts, including umbelliferous crops. In April 2023, parsley plantlets (Petroselinum crispum), showing stunted growth, yellowing, decayed roots and damping-off, were obtained from a nursery in central Slovenia, where parsley was grown in polystyrene trays in a greenhouse. Nearly 30% of plants were symptomatic. Sampled roots of ten plants contained ornamented oogonia (avg. 33.3 ± 1.4 µm in diam) with conical projections (5.2 ± 0.5 µm long) (Figure S1 A, B) in microscopically analyzed squash mounts. The pathogen was isolated from root pieces treated for surface disinfection with 0.5% sodium hypochlorite for 30 s, and washed with sterile water. Four 1-2 mm root pieces were taken from each of 10 plants, plated on the selective medium P5ARP, and incubated at 21 °C. Mycelia emerging from root pieces were transferred to carrot piece agar (CPA). Twenty-two equally looking oomycetous colonies were obtained; all sampled plants were infested. Oogonia formed by all colonies were similar to those observed on decayed roots and suggested that Globisporangium (Pythium) mastophorum is the causal disease agent. Analyses of partial ß-tubulin (Kroon et al. 2004) and mitochondrial cytochrome c oxidase I (COI) gene sequences (Robideau et al. 2011) confirmed the identification. Obtained COI (Genbank accession number OR725417) sequence was 100% identical to that from G. mastophorum strain CBS 375.72 (EU350523), whereas the ß-tubulin sequence (OR725416) corresponded to 99.6 % pairwise identity (KJ595502). Further, pathogenicity of an obtained isolate was tested on 4 wk-old curly leaf (cv. Petra F1) parsley. Half of a 7 d-old CPA culture, consisting of mycelium and oogonia, was finely cut and mixed with ca 50 ml of nonsterile commercial substrate (Potgrond H, AGRO-FertiCrop) in each of six 400 ml pots. Pots were filled with ca 300 ml additional substrate, into which 5 parsley seedlings were planted. Control plants were treated equally but with sterile CPA. Plantlets were watered with sterile tap water and held at ambient light conditions and temperature (night 18 °C - day 23 °C). After 14 d, inoculated plants started wilting and yellowing and showed stunted growth. After 21 d, roots were severely decayed and the seedlings damped-off (Figure S1 C). Four pieces each from 10 decayed roots were plated. Thirty-one pieces revealed pythium-like colonies. Obtained isolates were morphologically identical to the strain used for inoculation and identified as G. mastophorum. Control plants developed no foliar or root symptoms and no pythium-like species was obtained. Agricultural advisors observed occurrence of parsley damping-off also in other nurseries in Slovenia what may lead to spreading the pathogen to parsley in production fields and private gardens. The case emphasizes the need for implementing phytosanitary measures in order to eliminate primary inoculum. Reports from field-infected plants showed that G. mastophorum is a pathogen of parsley in Australia (Petkowski et al. 2013) and the USA (Tsuchida et al. 2018), and celery in the Czech Republic (Safránková and Holková 2017). Others isolated G. mastophorum from parsley in The Netherlands (online database of the Westerdijk Fungal Biodiversity Institute, strain CBS 243.86). However, the here described case is, to the best of our knowledge, one of the rare documentations of damping-off due to G. mastophorum in Europe (Safránková and Holková 2017) and the first in Slovenia. Funding: The work was funded by the Ministry of Agriculture, Forestry and Food of Slovenia, and Slovenian Research and Innovation Agency (ARIS Programs P4-0431 and P4-0072).

First report of Colletotrichum incanum causing leaf spots on common bean in Europe (Slovenia).

A Phaseolus vulgaris L. leaf showing necrotic spots was collected in an experimental bean field in central Slovenia in August 2021. The field contained diverse common bean lines sourced from genebank collections, with each line represented by 10 plants. While symptomatic leaves were seen across various lines, the reported species derived exclusively from a Huasca Huallaga Colorado plant (single-seed descent, USDA accession PI153714, doi: 10.18730/H7P9N), a Peruvian landrace. After incubating the leaf for 2 d at ambient temperature in a moist chamber, setose acervuli developed producing curved, distally tapering and proximately truncated conidia. Single-spore cultures developed equally-shaped conidia measuring 14.5-21.5 (avg. 18.5) × 3-4 (avg. 3.5) µm (n=60) on corn meal agar when mounted in lactic acid. Obtained morphological characters and sequences of the partial actin (GenBank accession, OR208162), beta-tubulin (OR208164), and histone 3 (OR208165) gene identified the isolate as Colletotrichum incanum H.-C. Yang, J.S. Haudenshield & G.L. Hartman. Sequences were identical to those from CBS 133485 (= NRRL 62592, IL6A), ex-type strain of C. incanum (KC110823, KC110814, and KC110796). Partial sequences of the chitin synthase (CHS) gene (OR208163), not available for the ex-type strain, was identical to sequences of other C. incanum strains reported from China (KP145539, ON189040, and OQ613679-OQ613686) or differed in two nucleotide positions (OL471268 and OL471269). The strain from Slovenia was deposited in the CBS biobanks of the Westerdijk Fungal Biodiversity Institute (Utrecht, The Netherlands) as CBS 150848. Pathogenicity of the strain was tested by spraying ca. 3×105 conidia as a watery spore suspension onto each leaf of 6 greenhouse-grown and 3 wk-old common bean plantlets (cv. KIS Amand). Nonsterile commercial substrate (Potgrond H, AGRO-FertiCrop) was used and natural light conditions at ambient temperatures (18-23°C) applied. Sterile water was sprayed on 6, equally grown negative control plants. Treated plants showed small brownish spots after 3 wks similar to those described by Yang et al. (2014) on soybean. Setose acervuli formed within 5 days after detached leaves were incubated in moist chambers. No acervuli formed on negative control plants. Conidia re-isolated from these acervuli and obtained cultures were morphologically identical to originally obtained conidia and cultures and those used for performing the pathogenicity test. Anthracnose is an important disease of common bean attributed to various races of C. lindemuthianum (Sacc. & Magnus) Briosi & Cavara (Nunes et al. 2021). Reporting an additional agent potentially able to cause diseases in common bean and so far not known to occur in Europe is of high relevance as the various genetic bean lines used in Europe may show alternative susceptibility levels to it. However, symptoms caused by C. incanum seem to be less severe as those caused by C. lindemuthianum and the species belongs to the C. spaethianum species complex, whose members have so far not been considered as pathogens of economic importance (Talhinhas & Baroncelli 2021). Yang et al. (2014) based C. incanum on isolates from soybean petioles (USA) and associated it with common bean by re-identifying strain ATCC 64682 obtained by Tu (1990) in Canada. Database queries revealed that it was encountered also on sugar beet (USA; Hanson et al. 2023) and on various crop hosts in China (e.g., chili; Diao et al. 2017), but not in Europe. The work was funded by the Ministry of Agriculture, Forestry and Food and conducted as part of research programs P4-0072 and P4-0431, financed by the Slovenian Research and Innovation Agency ARIS, and the Horizon 2020 project INCREASE funded by the European Union.

Comparative genomics highlights the importance of drug efflux transporters during evolution of mycoparasitism in Clonostachys subgenus Bionectria (Fungi, Ascomycota, Hypocreales).

Various strains of the mycoparasitic fungal species Clonostachys rosea are used commercially as biological control agents for the control of fungal plant diseases in agricultural crop production. Further improvements of the use and efficacy of C. rosea in biocontrol require a mechanistic understanding of the factors that determines the outcome of the interaction between C. rosea and plant pathogenic fungi. Here, we determined the genome sequences of 11 Clonostachys strains, representing five species in Clonostachys subgenus Bionectria, and performed a comparative genomic analysis with the aim to identify gene families evolving under selection for gene gains or losses. Several gene families predicted to encode proteins involved in biosynthesis of secondary metabolites, including polyketide synthases, nonribosomal peptide syntethases and cytochrome P450s, evolved under selection for gene gains (p ≤ .05) in the Bionectria subgenus lineage. This was accompanied with gene copy number increases (p ≤ .05) in ATP-binding cassette (ABC) transporters and major facilitator superfamily (MFS) transporters predicted to contribute to drug efflux. Most Clonostachys species were also characterized by high numbers of auxiliary activity (AA) family 9 lytic polysaccharide monooxygenases, AA3 glucose-methanol-choline oxidoreductases and additional carbohydrate-active enzyme gene families with putative activity (or binding) towards xylan and rhamnose/pectin substrates. Particular features of the C. rosea genome included expansions (p ≤ .05) of the ABC-B4 multidrug resistance transporters, the ABC-C5 multidrug resistance-related transporters and the 2.A.1.3 drug:H + antiporter-2 MFS drug resistance transporters. The ABC-G1 pleiotropic drug resistance transporter gene abcG6 in C. rosea was induced (p ≤ .009) by exposure to the antifungal Fusarium mycotoxin zearalenone (1121-fold) and various fungicides. Deletion of abcG6 resulted in mutants with reduced (p < .001) growth rates on media containing the fungicides boscalid, fenhexamid and iprodione. Our results emphasize the role of biosynthesis of, and protection against, secondary metabolites in Clonostachys subgenus Bionectria.

Bioaugmentation of Entomopathogenic Fungi for Sustainable Agriotes Larvae (Wireworms) Management in Maize.

Soil microorganisms influence biotic and abiotic stress tolerance of crops. Most interactions between plant symbiotic and non-symbiotic soil microorganisms and plants occur in the rhizosphere and are sustained through plant exudation/rhizodeposition. Bioaugmentation, i.e., the introduction or amplification of certain plant beneficial microbes (e.g., entomopathogenic fungi) into the rhizosphere, could contribute to controlling insect crop pests and replacing chemical, environmentally unfriendly insecticides. Wireworms, the soil-burrowing larval stages of click beetles (Coleoptera: Elateridae), are major pests of crops including maize, wheat and potatoes, worldwide. Alternative strategies for controlling wireworms are needed because several chemical pesticides used successfully in the past are being phased out because of their ecotoxicity. Therefore, virulence to Agriotes lineatus L. wireworms and plant beneficial traits of entomopathogenic fungi were investigated in a series of laboratory experiments. Tested taxa included environmentally retrieved Metarhizium brunneum Petch. (two strains), M. robertsii Bisch., Rehner & Humber (Hypocreales: Clavicipitaceae), and Beauveria brongniartii (Sacc.) Petch. and commercially formulated B. bassiana (Bals.-Criv.) Vuill. (Cordycipitaceae) and Bacillus thuringiensis Berliner 1915 var. kurstaki. In-house reared larvae were dipped in conidial suspension, and maize and wheat seeds were coated with fungal conidia. Metarhizium brunneum strains 1154 and 1868 significantly increased wireworm mortality. Fungi were significantly more often re-isolated from maize than wheat rhizoplanes in laboratory assays. The strains tested were rarely isolated as endophytes. Metarhizium brunneum strain 1154 stimulated wheat growth, while M. robertsii 1880 stimulated maize growth, whereas M. brunneum 1868 and others did not affect root or shoot length or plant biomass significantly in laboratory settings. Metarhizium brunneum strain 1868, re-isolated most often from maize rhizoplane, caused the highest wireworm mortality. It was further evaluated whether M. brunneum 1868 can protect maize varieties FeroXXY, LG 34.90 and Chapalu from wireworm damage and promote plant growth at field conditions. Plants of all three varieties stemming from seeds treated with conidia of M. brunneum 1868 showed significantly less wireworm damage 3 to 4 weeks after sowing (5- to 6-leaf stage) resulting in a significantly higher initial maize stand. However, only in the variety LG 34.90 a significant increase of the maize stand was observed at harvest time.

Out in the Cold: Identification of Genomic Regions Associated With Cold Tolerance in the Biocontrol Fungus Clonostachys rosea Through Genome-Wide Association Mapping.

There is an increasing importance for using biocontrol agents in combating plant diseases sustainably and in the long term. As large scale genomic sequencing becomes economically viable, the impact of single nucleotide polymorphisms (SNPs) on biocontrol-associated phenotypes can be easily studied across entire genomes of fungal populations. Here, we improved a previously reported genome assembly of the biocontrol fungus Clonostachys rosea strain IK726 using the PacBio sequencing platform, which resulted in a total genome size of 70.7 Mbp and 21,246 predicted genes. We further performed whole-genome re-sequencing of 52 additional C. rosea strains isolated globally using Illumina sequencing technology, in order to perform genome-wide association studies in conditions relevant for biocontrol activity. One such condition is the ability to grow at lower temperatures commonly encountered in cryic or frigid soils in temperate regions, as these will be prevalent for protecting growing crops in temperate climates. Growth rates at 10°C on potato dextrose agar of the 53 sequenced strains of C. rosea were measured and ranged between 0.066 and 0.413 mm/day. Performing a genome wide association study, a total of 1,478 SNP markers were significantly associated with the trait and located in 227 scaffolds, within or close to (< 1000 bp distance) 265 different genes. The predicted gene products included several chaperone proteins, membrane transporters, lipases, and proteins involved in chitin metabolism with possible roles in cold tolerance. The data reported in this study provides a foundation for future investigations into the genetic basis for cold tolerance in fungi, with important implications for biocontrol.

Effects of soil erosion on agro-ecosystem services and soil functions: A multidisciplinary study in nineteen organically farmed European and Turkish vineyards.

This multidisciplinary research work evaluated the effects of soil erosion on grape yield and quality and on different soil functions, namely water and nutrient supply, carbon sequestration, organic matter recycling, and soil biodiversity, with the aim to understand the causes of soil malfunctioning and work out a proper strategy of soil remediation. Degraded areas in nineteen organically farmed European and Turkish vineyards resulted in producing significantly lower amounts of grapes and excessive concentrations of sugar. Plants suffered from decreased water nutrition, due to shallower rooting depth, compaction, and reduced available water capacity, lower chemical fertility, as total nitrogen and cation exchange capacity, and higher concentration of carbonates. Carbon storage and organic matter recycling were also depressed. The general trend of soil enzyme activity mainly followed organic matter stock. Specific enzymatic activities suggested that in degraded soils, alongside a general slowdown in organic matter cycling, there was a greater reduction in decomposition capacity of the most recalcitrant forms. The abundance of Acari Oribatida and Collembola resulted the most sensitive indicator of soil degradation among the considered microarthropods. No clear difference in overall microbial richness and evenness were observed. All indices were relatively high and indicative of rich occurrence of many and rare microbial species. Dice cluster analyses indicated slight qualitative differences in Eubacterial and fungal community compositions in rhizosphere soil and roots in degraded soils. This multidisciplinary study indicates that the loss of soil fertility caused by excessive earth movement before planting, or accelerated erosion, mainly affects water nutrition and chemical fertility. Biological soil fertility is also reduced, in particular the ability of biota to decompose organic matter, while biodiversity is less affected, probably because of the organic management. Therefore, the restoration of the eroded soils requires site-specific and intensive treatments, including accurately chosen organic matrices for fertilization, privileging the most easily decomposable. Restoring soil fertility in depth, however, remain an open question, which needs further investigation.

Occultifur mephitis f.a., sp. nov. and other yeast species from hypoxic and elevated CO2 mofette environments.

An inventory of culturable yeasts from the soil and water of natural CO2 springs (mofettes) in northeast Slovenia is presented. In mofettes, CO2 of geological origin reaches the soil surface causing temporarily and spatially stable hypoxic environments in soil and water. In total, 142 yeast strains were isolated and identified from high CO2 and control meadow soil, meadow ground-water, forest pond and stream water. All water locations showed below-ground CO2 release. They were assigned to six basidiomycetous yeast genera (six species) and 11 ascomycetous genera (18 species). All ascomycetous yeasts, with the exception of Debaryomyces hansenii, were able to grow under elevated CO2 and fermented glucose. Candida sophiae-reginae, Pichia fermentans and Candida vartiovaarae were the dominating species in meadow and forest high CO2 exposed water. Meyerozyma guilliermondii and Wickerhamomyces anomalus predominated in high CO2 exposed soils. Using high dilution plating of a mofette soil sample, four strains of an unknown basidiomycetous species were isolated and are here newly described as Occultifur mephitis based on molecular phylogenetic and phenotypic criteria. The type strain of Occultifur mephitis is EXF-6436T[CBS 14611=PYCC 7049, LT594852 (D1/D2), KX929055 (ITS)]. An additional three isolated strains are EXF-6437 (LT594853, KX929056), EXF-6473 (LT594863, KX929057) and EXF-6482 (LT594867, KX929054), as well as a strain reported from previous studies isolated from a leaf of Cistus albidus in Portugal (CBS 10223=PYCC 6067), EU002842 (D1/D2), KY308183 (ITS).

Elsinopirins A-D, Decalin Polyketides from the Ascomycete Elsinoё pyri.

In course of our screening for new secondary metabolites from ecological niche specialized, phytopathogenic fungi, the plant pathogen Elsinoё pyri, strain 2203C, was found to produce four novel compounds (1-4), which were named elsinopirins A-D, in addition to the known metabolite elsinochrome A (5). After isolation by preparative high-performance liquid chromatography (HPLC), their structures, including relative stereochemistry, were elucidated by 1D and 2D nuclear magnetic resonance (NMR) and mass spectrometry (MS) data. Finally, absolute stereochemistry was assigned by chemical shifts of Mosher's esters (α-methoxy-α-trifluoromethylphenylacetic acid; MTPA) derivatives of elsinopirin B (2). The compounds were found to be devoid of significant antibacterial, antifungal, and cytotoxic activities.

Comparison of cauliflower-insect-fungus interactions and pesticides for cabbage root fly control.

Cabbage root fly (Delia radicum L.) control represents a major challenge in brassica production, therefore different management strategies for its control were tested in conventionally managed open field cauliflower production. Strategies included treatments with low-risk methods such as nitrogen lime, the insecticide spinosad and the Beauveria bassiana ATCC 74040-based biopesticide Naturalis. Their effects were compared with treatments based on nonformulated fungal species Metarhizium brunneum, B. bassiana, Clonostachys solani, Trichoderma atroviride, T. koningiopsis, and T. gamsii and commercial insecticides λ-cyhalothrin and thiamethoxam. Spinosad and thiamethoxam were pipetted to individual plants before transplanting; λ-cyhalothrin was sprayed after transplanting; nitrogen lime was applied at first hoeing. Nonformulated fungi were delivered onto cauliflower plantlets' roots as a single pretransplantation inoculation. The cabbage root fly population dynamics exhibited a strong spatiotemporal variation. The lowest number of cabbage root fly pupae recovered from cauliflower roots in the field experiments was recorded in plants treated with spinosad (significant reduction), followed by Naturalis and one of the tested M. brunneum strains (nonsignificant reduction). Significantly more pupae were counted in the nitrogen lime treatment. The field experiments showed that a single drench of cauliflower plantlets with spinosad offered consistent and enduring cabbage root fly control. Naturalis and nonformulated fungal isolates did not decrease cabbage root fly pressure significantly, apparently due to lack of statistical power. The implications of the substantial intra- and inter-annual pest pressure variation and the benefits of using single plant treatments are discussed, and recommendations for improvement of rhizosphere-competence utilizing biological control strategies provided.

Ijuhya vitellina sp. nov., a novel source for chaetoglobosin A, is a destructive parasite of the cereal cyst nematode Heterodera filipjevi.

Cyst nematodes are globally important pathogens in agriculture. Their sedentary lifestyle and long-term association with the roots of host plants render cyst nematodes especially good targets for attack by parasitic fungi. In this context fungi were specifically isolated from nematode eggs of the cereal cyst nematode Heterodera filipjevi. Here, Ijuhya vitellina (Ascomycota, Hypocreales, Bionectriaceae), encountered in wheat fields in Turkey, is newly described on the basis of phylogenetic analyses, morphological characters and life-style related inferences. The species destructively parasitises eggs inside cysts of H. filipjevi. The parasitism was reproduced in in vitro studies. Infected eggs were found to harbour microsclerotia produced by I. vitellina that resemble long-term survival structures also known from other ascomycetes. Microsclerotia were also formed by this species in pure cultures obtained from both, solitarily isolated infected eggs obtained from fields and artificially infected eggs. Hyphae penetrating the eggshell colonised the interior of eggs and became transformed into multicellular, chlamydospore-like structures that developed into microsclerotia. When isolated on artificial media, microsclerotia germinated to produce multiple emerging hyphae. The specific nature of morphological structures produced by I. vitellina inside nematode eggs is interpreted as a unique mode of interaction allowing long-term survival of the fungus inside nematode cysts that are known to survive periods of drought or other harsh environmental conditions. Generic classification of the new species is based on molecular phylogenetic inferences using five different gene regions. I. vitellina is the only species of the genus known to parasitise nematodes and produce microsclerotia. Metabolomic analyses revealed that within the Ijuhya species studied here, only I. vitellina produces chaetoglobosin A and its derivate 19-O-acetylchaetoglobosin A. Nematicidal and nematode-inhibiting activities of these compounds have been demonstrated suggesting that the production of these compounds may represent an adaptation to nematode parasitism.

Epitypification of Fusisporium (Fusarium) solani and its assignment to a common phylogenetic species in the Fusarium solani species complex.

Fusisporium solani was described as the causal agent of a dry rot of potato in Germany in the mid 19th century. As Fusarium solani, the species became known as a plurivorous plant pathogen, endophyte, decomposer, and opportunistic pathogen of humans and nutritional symbiont of insects. In parallel, it became evident that the morphologically defined species F. solani represents a phylogenetically and biologically complex group of often morphologically cryptic species that has come to be known in part as the F. solani species complex (FSSC), accommodating several formae speciales and mating populations/biological species. The FSSC currently includes more than 60 phylogenetic species. Several of these have been named, but the majority remains unnamed and the identity of F. solani sensu stricto is unclear. To promote further taxonomic developments in the FSSC, lectoand epitypification is proposed for Fusisporium solani Although no type material for F. solani is known to exist, the species was abundantly illustrated in the protologue. Thus, a relevant illustration provided by von Martius is selected as the lectotype. The epitype selected here originates from a rotting potato collected in a field in Slovenia. This strain causes a dry rot of artificially inoculated potatoes. It groups in the heretofore unnamed phylogenetic species 5, which is nested within clade 3 of the FSSC (FSSC 5). Members of this phylogenetic species have a wide geographic distribution and include soil saprotrophs and plant and opportunistic human pathogens. This typification is consistent with the original description of Fusisporium solani and the concept of F. solani as a widely distributed soil inhabitant and pathogen.

A Taxonomic Revision of the Wallemia sebi Species Complex.

Wallemia sebi is a xerophilic food- and air-borne fungus. The name has been used for strains that prevail in cold, temperate and tropical climates. In this study, multi-locus phylogenetic analyses, using the internal transcribed spacer (ITS) regions, DNA replication licensing factor (MCM7), pre-rRNA processing protein (TSR1), RNA polymerase II largest subunit (RPB1), RNA polymerase II second largest subunit (RPB2) and a new marker 3´-phosphoadenosine-5´-phosphatase (HAL2), confirmed the previous hypothesis that W. sebi presents a complex of at least four species. Here, we confirm and apply the phylogenetic analyses based species hypotheses from a companion study to guide phenotypic assessment of W. sebi like strains from a wide range of substrates, climates and continents allowed the recognition of W. sebi sensu stricto and three new species described as W. mellicola, W. Canadensis, and W. tropicalis. The species differ in their conidial size, xerotolerance, halotolerance, chaotolerance, growth temperature regimes, extracellular enzyme activity profiles, and secondary metabolite patterns. A key to all currently accepted Wallemia species is provided that allow their identification on the basis of physiological, micromorphological and culture characters.

Production of Obionin A and Derivatives by the Sooty Blotch Fungus Microcyclospora malicola.

A multitude of sooty blotch and flyspeck fungi, mainly belonging to the Ascomycetes order Capnodiales, causes dark blemishes and flyspeck-like spots on apples worldwide. Different sooty blotch and flyspeck fungi can coexist in the same orchard and even on a single fruit. Our preceding experiments revealed an activity of Microcyclospora malicola strain 1930 against the anthracnose fungus Colletotrichum fioriniae in dual culture assays. Extracts of M. malicola strain 1930 showed a broad bioactivity against filamentous fungus Mucor hiemalis and gram-positive bacterium Bacillus subtilis. A bioactivity-guided isolation led to the identification of obionin A (1) as the main active principle. In addition to 1, which was previously isolated from the marine fungus Leptosphaeria obiones, we isolated three derivatives. Metabolite 2 bears a keto function at C-6, besides the replacement of oxygen by nitrogen at position 10. Two more derivatives are adducts (3, 4) of acetone as work-up artifacts. Because obionin A (1) and its derivative 2 showed cytotoxic effects and antifungal activities, we propose a role of these secondary metabolites in the antagonism between M. malicola and other apple colonizing sooty blotch and flyspeck fungi, other epiphytes, or apple pathogens competing for the same ecological niche.

Candida and Fusarium species known as opportunistic human pathogens from customer-accessible parts of residential washing machines.

Energy constraints have altered consumer practice regarding the use of household washing machines. Washing machines were developed that use lower washing temperatures, smaller amounts of water and biodegradable detergents. These conditions may favour the enrichment of opportunistic human pathogenic fungi. We focused on the isolation of fungi from two user-accessible parts of washing machines that often contain microbial biofilms: drawers for detergents and rubber door seals. Out of 70 residential washing machines sampled in Slovenia, 79% were positive for fungi. In total, 72 strains belonging to 12 genera and 26 species were isolated. Among these, members of the Fusarium oxysporum and Fusarium solani species complexes, Candida parapsilosis and Exophiala phaeomuriformis represented 44% of fungi detected. These species are known as opportunistic human pathogens and can cause skin, nail or eye infections also in healthy humans. A machine learning analysis revealed that presence of detergents and softeners followed by washing temperature, represent most critical factors for fungal colonization. Three washing machines with persisting malodour that resulted in bad smelling laundry were analysed for the presence of fungi and bacteria. In these cases, fungi were isolated in low numbers (7.5 %), while bacteria Micrococcus luteus, Pseudomonas aeruginosa, and Sphingomonas species prevailed.

Evaluation of insect associated and plant growth promoting fungi in the control of cabbage root flies.

Delia radicum L. or cabbage maggot is an important pest for Brassicaceous crops. There are currently no registered chemical control agents for its control in Slovenia. Fungal control agents for cabbage maggot were therefore sought among nine rhizosphere-compatible and plant growth-promoting, soil-adapted, and entomopathogenic species to cabbage maggots and were assayed in in vitro and soil laboratory bioassays. In the in vitro tests, the conidial suspensions were applied directly to cabbage maggot eggs. The soil tests mimicked pathways of natural exposure of various insect life stages to the fungal strains. Conidial concentrations used in soil tests were comparable to economic rates for in-furrow application. The following fungi were tested: Trichoderma atroviride P. Karst. (2 isolates), Trichoderma koningiopsis Samuels, C. Suárez & H.C. Evans (1), Trichoderma gamsii Samuels & Druzhin. (3), Beauveria brongniartii (Saccardo) Petch (1), Beauveria bassiana (Balsamo-Crivelli) Vuillemin (2), Metarhizium robertsii J.F. Bisch., Rehner & Humber (1), Metarhizium anisopliae (Metschn.) Sorokin (4), Purpureocillium lilacinum (Thom) Luangsa-ard, Houbraken, Hywel-Jones & Samson (2), and Clonostachys solani f. nigrovirens (J.F.H. Beyma) Schroers (2). Abbott's corrected mortality in the in vitro tests ranged from 0.0 +/- 18.9 to 47.6 +/- 9.0% and in the soil test from 2.4 +/- 13.0 to 68.2 +/- 21.5%. Seven isolates (B. bassiana [isolate 1174], C. solani [1828], M. anisopliae [1154 and 1868], T. atroviride [1872], T. koningiopsis [1874], and T. gamsii [1876]) caused significant cabbage maggot mortality in either in vitro or soil tests. The importance of fungal ecology as a criterion during the screening of potential biological control agents is discussed.

Direct plantlet inoculation with soil or insect-associated fungi may control cabbage root fly maggots.

A potential Delia radicum biological control strategy involving cauliflower plantlet inoculation with various fungi was investigated in a series of laboratory and glasshouse experiments. In addition to entomopathogenic fungi, fungi with a high rhizosphere competence and fungi with the ability to survive as saprotrophs in soil were tested. The following fungal species were evaluated in the experiments: Trichoderma atroviride, T. koningiopsis, T. gamsii, Beauveria bassiana, Metharhizium anisopliae, M. brunneum and Clonostachys solani. A commercial carbosulfan-based insecticide was used as a positive control. Additionally, two commercial products, one based on B. bassiana (Naturalis) and one on Bacillus thuringiensis (Delfin) were used as reference biocontrol agents. The aims were (i) to assess the pathogenicity of the selected fungal isolates to Delia radicum, (ii) to evaluate the fungal isolates' rhizosphere competence, with the emphasis on the persistence of the original inoculum on the growing roots, (iii) to assess possible endophytic plant tissue colonization, and (iv) to evaluate potential plant growth stimulating effects of the added inoculi. Significant pathogenicity of tested fungi against Delia radicum was confirmed in in vitro and glasshouse experiments. All tested fungi persisted on cauliflower rhizoplane. More importantly, the added fungi were found on thoroughly washed roots outside the original point of inoculation. This provided us with evidence that our tested fungi could be transferred via or grow with the elongating roots. In addition to colonizing the rhizoplane, some fungi were found inside the plant root or stem tissue, thus exhibiting endophytic characteristics. The importance of fungal ecology as a criterion in appropriate biological control agent selection is discussed.

Peltaster cerophilus is a new species of the apple sooty blotch complex from Europe.

Adopting the currently used concept for the genus Peltaster, the sooty blotch fungus Peltaster cerophilus is newly described from the cuticle of ripening or ripe apples. It forms a punctate phenotype consisting of superficially formed pycnothyria and a superficial mycelial mat consisting of a net of brown or brownish black hyphae. The pycnothyria are olivaceous brown to brown but have a spot in the center that is less strongly pigmented. Pycnothyria on the holotype of P. fructicola are homogeneously pigmented. On synthetic nutrient-poor agar, P. cerophilus is largely indistinguishable from P. fructicola. It forms delicate, spreading hyphae and intercalary conidiogenous cells with short, lateral, apically thick-walled conidiogenous necks forming blastic, unpigmented, one-celled conidia in basipetal succession. Conidia can swell and become one-septate. The species has microcyclical conidiation in proximate parts of colonies. DNA sequence analyses based on the ITS and the partial nuclear small and large subunit ribosomal RNA genes, the partial mitochondrial small subunit rRNA gene and the partial translation elongation factor 1-α gene support the distinction of the European P. cerophilus from P. fructicola, which is known from North America and Europe. The nuclear small ribosomal RNA subunit gene sequences of P. cerophilus contain two group I introns at locations known to accommodate introns in certain other, unrelated taxa. One of these, for which the code "SSU-1506 intron" was adopted, is 1459 base pairs long and located between the universal primer sites ITS5 and ITS1. Similar or positional-differing introns were encountered also in three currently undescribed Peltaster species. Representative strains of Peltaster fructicola did not accommodate introns in the nuclear small subunit ribosomal RNA gene.

Production of Trichothecenes by the apple sooty blotch fungus Microcyclospora tardicrescens.

The sooty blotch and flyspeck (SBFS) syndrome of apples and other fruits is caused by a complex consortium of epiphytic fungi that colonize the fruit cuticula. SBFS fungal strains isolated from apples were screened for growth inhibition of the phytopathogen Colletotrichum fioriniae in dual culture tests. Extracts of 11 isolates of SBFS fungi (Microcyclospora malicola, Microcyclospora pomicola, Microcyclospora tardicrescens, and Microcyclosporella mali) inhibited growth of the test strains and were studied for production of antibiotics. A strain of Microcyclospora tardicrescens strongly inhibited growth and was cultivated on a larger scale to characterize its secondary metabolites. Bioassay-guided fractionation and subsequent structure elucidation by spectroscopic and spectrometric methods (NMR, HRMS) yielded trichothecolone acetate (1) and its novel derivative (S)-7-hydroxytrichothecolone acetate (2) as active principles. Microcyclospora tardicrescens was thus identified as a producer of the hazardous trichothecene type mycotoxins for the first time, which should give reason to monitor these foodborne fungi more carefully in the future.

Genera in Bionectriaceae, Hypocreaceae, and Nectriaceae (Hypocreales) proposed for acceptance or rejection.

With the recent changes concerning pleomorphic fungi in the new International Code of Nomenclature for algae, fungi, and plants (ICN), it is necessary to propose the acceptance or protection of sexual morph-typified or asexual morph-typified generic names that do not have priority, or to propose the rejection or suppression of competing names. In addition, sexual morph-typified generic names, where widely used, must be proposed for rejection or suppression in favour of asexual morph-typified names that have priority, or the latter must be proposed for conservation or protection. Some pragmatic criteria used for deciding the acceptance or rejection of generic names include: the number of name changes required when one generic name is used over another, the clarity of the generic concept, their relative frequencies of use in the scientific literature, and a vote of interested mycologists. Here, twelve widely used generic names in three families of Hypocreales are proposed for acceptance, either by conservation or protection, despite their lack of priority of publication, or because they are widely used asexual morph-typified names. Each pair of generic names is evaluated, with a recommendation as to the generic name to be used, and safeguarded, either through conservation or protection. Four generic names typified by a species with a sexual morph as type that are younger than competing generic names typified by a species with an asexual morph type, are proposed for use. Eight older generic names typified by species with an asexual morph as type are proposed for use over younger competing generic names typified by a species with a sexual morph as type. Within Bionectriaceae, Clonostachys is recommended over Bionectria; in Hypocreaceae, Hypomyces is recommended over Cladobotryum, Sphaerostilbella over Gliocladium, and Trichoderma over Hypocrea; and in Nectriaceae, Actinostilbe is recommended over Lanatonectria, Cylindrocladiella over Nectricladiella, Fusarium over Gibberella, Gliocephalotrichum over Leuconectria, Gliocladiopsis over Glionectria, Nalanthamala over Rubrinectria, Nectria over Tubercularia, and Neonectria over Cylindrocarpon.

Internet-accessible DNA sequence database for identifying fusaria from human and animal infections.

Because less than one-third of clinically relevant fusaria can be accurately identified to species level using phenotypic data (i.e., morphological species recognition), we constructed a three-locus DNA sequence database to facilitate molecular identification of the 69 Fusarium species associated with human or animal mycoses encountered in clinical microbiology laboratories. The database comprises partial sequences from three nuclear genes: translation elongation factor 1α (EF-1α), the largest subunit of RNA polymerase (RPB1), and the second largest subunit of RNA polymerase (RPB2). These three gene fragments can be amplified by PCR and sequenced using primers that are conserved across the phylogenetic breadth of Fusarium. Phylogenetic analyses of the combined data set reveal that, with the exception of two monotypic lineages, all clinically relevant fusaria are nested in one of eight variously sized and strongly supported species complexes. The monophyletic lineages have been named informally to facilitate communication of an isolate's clade membership and genetic diversity. To identify isolates to the species included within the database, partial DNA sequence data from one or more of the three genes can be used as a BLAST query against the database which is Web accessible at FUSARIUM-ID (http://isolate.fusariumdb.org) and the Centraalbureau voor Schimmelcultures (CBS-KNAW) Fungal Biodiversity Center (http://www.cbs.knaw.nl/fusarium). Alternatively, isolates can be identified via phylogenetic analysis by adding sequences of unknowns to the DNA sequence alignment, which can be downloaded from the two aforementioned websites. The utility of this database should increase significantly as members of the clinical microbiology community deposit in internationally accessible culture collections (e.g., CBS-KNAW or the Fusarium Research Center) cultures of novel mycosis-associated fusaria, along with associated, corrected sequence chromatograms and data, so that the sequence results can be verified and isolates are made available for future study.