Jan Dijksterhuis

Germination of Penicillium paneum Conidia Is Regulated by 1-Octen-3-ol, a Volatile Self-Inhibitor

ABSTRACT Penicillium paneum is an important contaminant of cereal grains which is able to grow at low temperature, low pH, high levels of carbon dioxide, and under acid conditions. P. paneum produces mycotoxins, which may be harmful to animals and humans. We found that conidia in dense suspensions showed poor germination, suggesting the presence of a self-inhibitor. A volatile compound(s) produced by these high-density conditions also inhibited mycelial growth of different species of fungi belonging to a variety of genera, suggesting a broad action range. The heat-stable compound was isolated by successive centrifugation of the supernatant obtained from spore suspensions with a density of 109 conidia ml−1. By using static headspace analyses, two major peaks were distinguished, with the highest production of these metabolites after 22 h of incubation at 25°C and shaking at 140 rpm. Gas chromatography coupled with mass spectra analysis revealed the compounds to be 3-octanone and 1-octen-3-ol. Notably, only the latter compound appeared to block the germination process at different developmental stages of the conidia (swelling and germ tube formation). In this study, 1-octen-3-ol influenced different developmental processes during the P. paneum life cycle, including induction of microcycle conidiation and inhibition of spore germination. Therefore, the compound can be considered a fungal hormone during fungal development.

Food mycology : a multifaceted approach to fungi and food

Fungi and Living Crops Cross-talk Between Host and Fungus in Postharvest Situations and its Effect on Symptom Development, D. Prusky and P. Kolattukudy Real Time Monitoring of Ethylene during Fungal-Plant Interaction by Laser-Based Photoacoustic Spectroscopy, S.M. Cristescu, E.J. Woltering and F.J.M. Harren The Fungal Spore in Food Mycology Spore Formation in Food Relevant Fungi, U. Ugalde and L.M. Corrochano Dispersal of Fungal Spores through the Air, A. McCartney and J. West The Germinating Spore as a Contaminating Vehicle, G.S. Chitarra and J. Dijksterhuis Heat-Resistant Ascospores, J. Dijksterhuis Fungi and Mycotoxins Why do Fungi Produce Mycotoxins?, N. Magan and D. Aldred Mycotoxin Producers, J.C. Frisvad, U. Thrane and R.A. Samson Fungi as Hyperproducers Filamentous Fungi as Cell Factories for Metabolite Production, W. de Jongh and J. Nielsen Hyperproduction of Enzymes by Fungi, H.A.B. Wosten, K. Scholtmeijer and R.P. de Vries Fungal Spoilage Ecology, Growth and Detection Association of Moulds to Foods, J.C. Frisvad, B. Andersen and R.A. Samson Transport Phenomena in Fungal Colonisation on a Food Matrix, Y.S.P. Rahardjo and A. Rinzema Molecular Detection and Monitoring, R. Geisen Fungal Volatiles: Biomarkers of Good and Bad Food Quality, K. Karlshoj, Per V. Nielsen and T. Ostenfeld Larsen Wine and Fungi - Implications of Vineyard Infections, S.L. Leong Cheese and Fermented Sausages, J. Stark Fungi as Food The Colonizing Fungus as a Food Provider, R. Nout Fungal Protein for Food, U. Thrane Edible Mushrooms: From Industrial Cultivation to Collection from the Wild, J. Baar, G. Straatsma, I. Paradi and J.G.M. Amsing Index

Functionality and prevalence of trehalose-based oligosaccharides as novel compatible solutes in ascospores of Neosartorya fischeri (Aspergillus fischeri) and other fungi

Ascospores of Neosartorya, Byssochlamys and Talaromyces can be regarded as the most stress-resistant eukaryotic cells. They can survive exposure at temperatures as high as 85°C for 100 min or more. Neosartorya fischeri ascospores are more viscous and more resistant to the combined stress of heat and desiccation than the ascospores of Talaromyces macrosporus which contain predominantly trehalose. These ascospores contain trehalose-based oligosaccharides (TOS) that are novel compatible solutes, which are accumulated to high levels. These compounds are also found in other members of the genus Neosartorya and in some other genera within the order Eurotiales that also include Byssochlamys and Talaromyces. The presence of oligosaccharides was observed in species that had a relatively high growth temperature. TOS glasses have a higher glass transition temperature (Tg) than trehalose, and they form a stable glass with crystallizing molecules, such as mannitol. Our data indicate that TOS are important for prolonged stabilization of cells against stress. The possible unique role of these solutes in protection against dry heat conditions is discussed.

A decrease in bulk water and mannitol and accumulation of trehalose and trehalose-based oligosaccharides define a two-stage maturation process towards extreme stress resistance in ascospores of Neosartorya fischeri (Aspergillus fischeri).

Fungal propagules survive stresses better than vegetative cells. Neosartorya fischeri, an Aspergillus teleomorph, forms ascospores that survive high temperatures or drying followed by heat. Not much is known about maturation and development of extreme stress resistance in fungal cells. This study provides a novel two-step model for the acquisition of extreme stress resistance and entry into dormancy. Ascospores of 11- and 15-day-old cultures exhibited heat resistance, physiological activity, accumulation of compatible solutes and a steep increase in cytoplasmic viscosity. Electron spin resonance spectroscopy indicated that this stage is associated with the removal of bulk water and an increase of chemical stability. Older ascospores from 15- to 50-day-old cultures showed no changes in compatible solute content and cytoplasmic viscosity, but did exhibit a further increase of heat resistance and redox stability with age. This stage was also characterized by changes in the composition of the mixture of compatible solutes. Mannitol levels decreased and the relative quantities of trehalose and trehalose-based oligosaccharides increased. Dormant ascospores of N. fischeri survive in low-water habitats. After activation of the germination process, the stress resistance decreases, compatible solutes are degraded and the cellular viscosity drops. After 5 h, the hydrated cells enter the vegetative stage and redox stability has decreased notably.

High-oxygen and high-carbon dioxide containing atmospheres inhibit growth of food associated moulds.

Aims: The objective of this study was to determine the relationship between the growth of three foodborne fungi and high‐oxygen modified atmosphere.

Aspergillus penicillioides differentiation and cell division at 0.585 water activity

Water availability acts as the most stringent constraint for life on Earth. Thus, understanding the water relations of microbial extremophiles is imperative to our ability to increase agricultural productivity (e.g., by enhancing the processing and turnover of dead organic matter in soils of arid regions), reduce human exposure to mycotoxins in buildings and our food-supply chain, prevent the spoilage of foods/animal feeds, books, museum specimens and artworks and better control microbiology of industrial fermentations. Only a small number of microbial systems can retain activity at <0.710 water activity (ISME J 2015 9: 1333-1351). It has long-been considered that the most resilient of these is Xeromyces bisporus, which inhabits sugar-rich substrates (Appl Environ Microbiol 1968 16: 1853-1858). The current study focused on germination of Aspergillus penicillioides, a xerophile which is also able to grow under low humidity and saline conditions. Investigations of germination differed from those reported earlier: firstly, aerially borne conidia were harvested, and then used for inoculations, in their dry condition; secondly, cultures were incubated at 24°C, i.e. below optimum germination temperature, to minimize the possibility of water loss from the substrate; thirdly, cultures remained sealed throughout the 73-day study period (microscopic examination was carried out directly 48 through the Petri plate lid); fourthly, the germination parameters determined were: rates and extent of conidial swelling, production of differentiated germination-structures and septate germlings, and subsequent development of mycelium and/or sporulation; fifthly, assessments were carried out over a range of water-activity values and time points to obtain a complete profile of the germination process. Conidia swelled, formed differentiated germination-structures and then produced septate germlings at a water-activity of just 0.585 (≡58.5% relative humidity), outside the currently understood thermodynamic window for life. Furthermore, analyses of these data suggest a theoretical water-activity minimum of 0.565 for germination of A. penicilliodes. In relation to astrobiology, these findings have an application in understanding the limits to life in extraterrestrial environments. In light of current plans for exploration missions to Mars and other places, and the need to safeguard martian scientific sites and potential resources (including water) for future human habitation, a knowledge-based and effective policy for planetary protection is essential. As it is, Mars-bound spacecraft may frequently be contaminated with aspergilli (including A. penicillioides) and other organisms which, when transported to other planetary bodies, pose a contamination risk. In crafting countermeasures to offset this, it is important to know as precisely as possible the capabilities of these potential interplanetary visitors.

Hoffmannoscypha, a novel genus of brightly coloured, cupulate Pyronemataceae closely related to Tricharina and Geopora

The rare apothecial, cupulate fungus Geopora pellita (Pyronemataceae) is characterized by a uniquely bright yellow-orange excipulum. We here re-examine its affiliations by use of morphological, molecular phylogenetic and ultrastructural analyses. G. pellita appears as phylogenetically rather isolated, being the sister group of a clade comprising Phaeangium, Picoa, the majority of the Tricharina species, and the remaining Geopora species. Based on its phylogenetic position and its unique combination of morphological characters, we assign G. pellita to Hoffmannoscypha, gen. nov., as H. pellita, comb. nov. As in a previous study, analyses of both large subunit (LSU) and internal transcribed spacer (ITS) ribosomal DNA suggest that the remaining genus Geopora is paraphyletic, with the hypogeous, ptychothecial type species more closely related to Picoa and Phaeangium than to the greyish-brownish cupulate and apothecial Geopora spp., indicating that the latter should be reassigned to the genus Sepultaria. The current study also shows that ITS confirm LSU data regarding the polyphyly of Tricharina.

Separate effects of moisture content and water activity on the hyphal extension of Penicillium rubens on porous media

To prevent indoor fungal growth, understanding the moisture relations of fungi is a key element. Indoor moisture is quantified by the relative humidity (RH). RH controls the water activity of the indoor materials that fungi grow on, a well-studied parameter known to limit fungal growth. RH, however, also controls the amount of water present in these materials, the moisture content. The significance of the moisture content of these materials to indoor fungal growth is currently overlooked. In the work reported here, growth experiments with the indoor fungus Penicillium rubens on gypsum substrates were performed to test whether the moisture content influences growth on porous materials. Second, we report the development of a video microscopy method that for the first time quantified hyphal growth on a porous material. It is found that a higher moisture content leads to earlier colonization and higher hyphal extension rates. This is a fundamental step in unravelling the effect of RH on indoor fungal growth. The real-time monitoring of colonization of gypsum provides a new view of growth on indoor surfaces.

Water-, pH- and temperature relations of germination for the extreme xerophiles Xeromyces bisporus (FRR 0025), Aspergillus penicillioides (JH06THJ) and Eurotium halophilicum (FRR 2471).

Water activity, temperature and pH are determinants for biotic activity of cellular systems, biosphere function and, indeed, for all life processes. This study was carried out at high concentrations of glycerol, which concurrently reduces water activity and acts as a stress protectant, to characterize the biophysical capabilities of the most extremely xerophilic organisms known. These were the fungal xerophiles: Xeromyces bisporus (FRR 0025), Aspergillus penicillioides (JH06THJ) and Eurotium halophilicum (FRR 2471). High‐glycerol spores were produced and germination was determined using 38 media in the 0.995–0.637 water activity range, 33 media in the 2.80–9.80 pH range and 10 incubation temperatures, from 2 to 50°C. Water activity was modified by supplementing media with glycerol+sucrose, glycerol+NaCl and glycerol+NaCl+sucrose which are known to be biologically permissive for X. bisporus, A. penicillioides and E. halophilicum respectively. The windows and rates for spore germination were quantified for water activity, pH and temperature; symmetry/asymmetry of the germination profiles were then determined in relation to supra‐ and sub‐optimal conditions; and pH‐ and temperature optima for extreme xerophilicity were quantified. The windows for spore germination were ~1 to 0.637 water activity, pH 2.80–9.80 and > 10 and < 44°C, depending on strain. Germination profiles in relation to water activity and temperature were asymmetrical because conditions known to entropically disorder cellular macromolecules, i.e. supra‐optimal water activity and high temperatures, were severely inhibitory. Implications of these processes were considered in relation to the in‐situ ecology of extreme conditions and environments; the study also raises a number of unanswered questions which suggest the need for new lines of experimentation.

Nutrient limitation leads to penetrative growth into agar and affects aroma formation in Pichia fabianii, P. kudriavzevii and Saccharomyces cerevisiae

Among fermentative yeast species, Saccharomyces cerevisiae is most frequently used as a model organism, although other yeast species may have special features that make them interesting candidates to apply in food‐fermentation processes. In this study, we used three yeast species isolated from fermented masau (Ziziphus mauritiana) fruit, S. cerevisiae 131, Pichia fabianii 65 and Pichia kudriavzevii 129, and determined the impact of nitrogen and/or glucose limitation on surface growth mode and the production of volatile organic compounds (VOCs). All three species displayed significant changes in growth mode in all nutrient‐limited conditions, signified by the formation of metafilaments or pseudohyphae. The timing of the transition was found to be species‐specific. Transition in growth mode is suggested to be linked to the production of certain fusel alcohols, such as phenylethyl alcohol, which serve as quorum‐sensing molecules. Interestingly, we did not observe concomitant increased production of phenylethyl alcohol and filamentous growth. Notably, a broader range of esters was found only for the Pichia spp. grown on nitrogen‐limited agar for 21 days compared to nutrient‐rich agar, and when grown on glucose‐ and glucose‐ plus nitrogen‐limited agar. Our data suggest that for the Pichia spp., the formation of esters may play an important role in the switch in growth mode upon nitrogen limitation. Further biological or ecological implications of ester formation are discussed. Copyright