Toxin-producing endosymbionts shield pathogenic fungus against micropredators

The phytopathogenic fungus Rhizopus microsporus harbours a bacterial endosymbiont (Mycetohabitans rhizoxinica) for the production of the toxin rhizoxin, the causative agent of rice seedling blight. This toxinogenic bacterial-fungal alliance is, however, not restricted to the plant disease, but has been detected in numerous environmental isolates from geographically distinct sites covering all five continents. Yet, the ecological role of rhizoxin beyond rice seedling blight has been unknown. Here we show that rhizoxin serves the fungal host in fending off protozoan and metazoan predators. Fluorescence microscopy and co-culture experiments with the fungivorous amoeba Protostelium aurantium revealed that ingestion of R. microsporus spores is toxic to P. aurantium. This amoebicidal effect is caused by the bacterial rhizoxin congener rhizoxin S2, which is also lethal towards the model nematode Caenorhabditis elegans. By combining stereomicroscopy, automated image analyses, and quantification of nematode movement we show that the fungivorous nematode Aphelenchus avenae actively feeds on R. microsporus that is lacking endosymbionts, while worms co-incubated with symbiotic R. microsporus are significantly less lively. This work uncovers an unexpected ecological role of rhizoxin as shield against micropredators. This finding suggests that predators may function an evolutionary driving force to maintain toxin-producing endosymbionts in non-pathogenic fungi.

Reevaluating the Species Status of the Southern Ghost Pipe, Monotropa brittonii (Ericaceae)

Abstract— Relationships among members of Ericaceae subfamily Monotropoideae have been difficult to resolve due to reduction and convergent evolution in these parasitic plants. All species in this subfamily are fully mycoheterotrophic, meaning they obtain nutrients by parasitizing fungi rather than through photosynthesis. Here, we examine relationships and host specificity in one of the most widespread species in this subfamily, Monotropa uniflora. We use several lines of evidence to investigate whether there is support for recognizing a segregate, M. brittonii, as distinct. Based on molecular and morphological analysis of Monotropa collected throughout its range in the United States, we find two distinct lineages, one of which corresponds morphologically and geographically to Small’s M. brittonii. We identified several morphological characters that differ between the two species. We also observed a high degree of fungal host specificity in M. brittonii, which appears to parasitize almost exclusively Lactifluus subgenus Lactariopsis section Albati. Additionally, M. brittonii was primarily collected from Florida scrub, which are xeric, shrub-dominated habitats that differ substantially from the mesic forests where M. uniflora typically occurs. Based on these molecular, morphological, and ecological differences, we support recognition of M. brittonii as distinct from M. uniflora.

Pathogenic potential prediction for novel fungal DNA based on a curated fungi-hosts data collection

Background: Emerging pathogens are a growing threat, but large data collections and approaches for predicting the risk associated with novel agents are limited to bacteria and viruses. Pathogenic fungi, which also pose a constant threat to public health, remain understudied. Relevant, curated data remains comparatively scarce and scattered among many different sources, hindering the development of sequencing-based detection workflows for novel fungal pathogens. No prediction method working for agents across all three groups is available, even though the cause of an infection is often difficult to identify from symptoms alone. Results: We present a curated collection of fungal host range data, comprising records on human, animal and plant pathogens, as well as other plant-associated fungi, linked to publicly available genomes. We show that the resulting database can be used to predict the pathogenic potential of novel fungal species directly from DNA sequences with either sequence homology or deep learning. We develop learned, numerical representations of the collected genomes and show that the human pathogens are separable from non-human pathogens. Finally, we train multi-class models predicting if next-generation sequencing reads originate from novel fungal, bacterial or viral threats. Conclusions: The presented data collection enables accurate detection of novel pathogens from sequencing data. It is also a comprehensive resource that can find use beyond this particular task. This can include possible applications in proteomics and genomics, employing both machine learning and direct sequence comparison. Availability: The database and models are hosted at https://zenodo.org/record/5711852 and https://zenodo.org/record/5711877. Source code is available at https://gitlab.com/dacs-hpi/deepac.

Trichoderma reesei Contains a Biosynthetic Gene Cluster That Encodes the Antifungal Agent Ilicicolin H

The trili biosynthetic gene cluster (BGC) from the well-studied organism Trichoderma reesei was studied by heterologous expression in the fungal host Aspergillus oryzae. Coexpression of triliA and triliB produces two new acyl tetramic acids. Addition of the ring-expanding cytochrome P450 encoded by triliC then yields a known pyridone intermediate to ilicicolin H and a new chain-truncated shunt metabolite. Finally, addition of the intramolecular Diels-Alderase encoded by triliD affords a mixture of 8-epi ilicicolin H and ilicicolin H itself, showing that the T. reesei trili BGC encodes biosynthesis of this potent antifungal agent. Unexpected A. oryzae shunt pathways are responsible for the production of the new compounds, emphasising the role of fungal hosts in catalysing diversification reactions.

Transcriptional profiles of a foliar fungal endophyte (Pestalotiopsis, Ascomycota) and its endohyphal bacterium (Luteibacter, Gammaproteobacteria) in co-culture support sulfur exchange and growth regulation

Symbiosis with bacteria is widespread among eukaryotes, including fungi. Bacteria that live within fungal mycelia (endohyphal bacteria) occur in many plant-associated fungi, including diverse Mucoromycota and Dikarya. Pestalotiopsis sp. 9143 is a filamentous ascomycete isolated originally as a foliar endophyte of Platycladus orientalis (Cupressaceae). It is infected naturally with the endohyphal bacterium Luteibacter sp. 9143, which influences auxin and enzyme production by its fungal host. Previous studies have used transcriptomics to examine similar symbioses between endohyphal bacteria and root-associated fungi such as arbuscular mycorrhizal fungi and plant pathogens. However, currently there are no gene expression studies of endohyphal bacteria of Ascomycota, the most species-rich fungal phylum. We developed methods for assessing gene expression by Pestalotiopsis sp. and Luteibacter sp. when grown in co-culture and when each was grown axenically. Our assays showed that the density of Luteibacter sp. in co-culture was greater than in axenic culture, but the opposite was true for the Pestalotiopsis sp. Dual RNA-seq data demonstrate that growing in co-culture modulates developmental and metabolic processes in both the fungus and bacterium, potentially through changes in the balance of organic sulfur via methionine acquisition. Our analyses also suggest an unexpected, potential role of the bacterial type VI secretion system in symbiosis establishment, expanding current understanding of the scope and dynamics of fungal-bacterial symbioses.

Inflammasome-mediated GSDMD activation facilitates escape of Candida albicans from macrophages

Candida albicans is the most common cause of fungal sepsis. Inhibition of inflammasome activity confers resistance to polymicrobial and LPS-induced sepsis; however, inflammasome signaling appears to protect against C. albicans infection, so inflammasome inhibitors are not clinically useful for candidiasis. Here we show disruption of GSDMD, a known inflammasome target and key pyroptotic cell death mediator, paradoxically alleviates candidiasis, improving outcomes and survival of Candida-infected mice. Mechanistically, C. albicans hijacked the canonical inflammasome-GSDMD axis-mediated pyroptosis to promote their escape from macrophages, deploying hyphae and candidalysin, a pore-forming toxin expressed by hyphae. GSDMD inhibition alleviated candidiasis by preventing C. albicans escape from macrophages while maintaining inflammasome-dependent but GSDMD-independent IL-1β production for anti-fungal host defenses. This study demonstrates key functions for GSDMD in Candida’s escape from host immunity in vitro and in vivo and suggests that GSDMD may be a potential therapeutic target in C. albicans-induced sepsis.

Arbuscular Mycorrhizal Fungi in Agriculture

Arbuscular mycorrhizal (AM) fungi are biotrophic symbionts forming close relationships with an estimated 80% of terrestrial plants suitable as their host. Via an established AM fungal–host relationship, soil-bound nutrients are made available to the host plant through root cortical arbuscules as the site of exchange. At these sites, photosynthetic carbohydrates are provided to the AM fungus—carbohydrates that cannot be produced by the fungus. AM fungal–host symbiosis is very sensitive to soil disturbance, for example, agricultural tillage practices can damage and reduce AM fungal abilities to interact with a host and provide plant growth-promoting properties.

Diversity and Host Relationships of the Mycoparasite Sepedonium (Hypocreales, Ascomycota) in Temperate Central Chile

We present the first major survey of regional diversity, distribution and host-association of Sepedonium. Whereas the rather scarce worldwide records of this mycoparasitic fungus suggested no specific distribution pattern of most species before, we provide new evidence of endemic and specific host-parasite guilds of Sepedonium in Southern South America, including the description of a new species. The corresponding inventory was performed in temperate central Chile. The regional landscape, a mosaic of exotic timber plantations and remnants of native Nothofagus forests, facilitates a unique combination of endemic and adventitious Boletales hosts. During a two-year survey, 35 Sepedonium strains were isolated and cultured from infected basidiomata of allochthonous Chalciporus piperatus, Paxillus involutus, Rhizopogon spp. and Suillus spp., as well as from the native Boletus loyita, B. loyo, B. putidus and Gastroboletus valdivianus. Taxonomic diagnosis included morphology of conidia and conidiophores, sequences of ITS, RPB2 and EF1 molecular markers and characteristics of in vitro cultures. Phylogenetic reconstructions were performed using Bayesian methods. Four Sepedonium species could be identified and characterized, viz.: S. ampullosporum, S. chrysospermum, S. laevigatum and the newly described species S. loyorum. The most frequent species on introduced Boletales was S. ampullosporum, followed by S. chrysospermum and S. laevigatum. S. loyorum sp. nov. was found exclusively on native boletacean hosts, separated from its closest relative S. chalcipori by micromorphological and molecular attributes. Species descriptions and identification keys are provided. Ecological and biogeographical aspects of endemic and allochthonous symbiotic units consisting of mycoparasite, ectomycorrhizal fungal host and respective mycorrhizal tree are discussed.

Identifying fungal-host associations in an amphibian host system

Host-associated microbes can interact with macro-organisms in a number of ways that affect host health. Few studies of host-associated microbiomes, however, focus on fungi. In addition, it is difficult to discern whether a fungal organism found in or on an ectotherm host is associating with it in a durable, symbiotic interaction versus a transient one, and to what extent the habitat and host share microbes. We seek to identify these host-microbe interactions on an amphibian, the Colorado boreal toad (Anaxyrus boreas boreas). We sequenced the ITS1 region of the fungal community on the skin of wild toads (n = 124) from four sites in the Colorado Rocky Mountains, across its physiologically dynamic developmental life stages. We also sampled the common habitats used by boreal toads: water from their natal wetland and aquatic pond sediment. We then examined diversity patterns within different life stages, between host and habitat, and identified fungal taxa that could be putatively host-associated with toads by using an indicator species analysis on toad versus environmental samples. Host and habitat were strikingly similar, with the exception of toad eggs. Post-hatching toad life stages were distinct in their various fungal diversity measures. We identified eight fungal taxa that were significantly associated with eggs, but no other fungal taxa were associated with other toad life stages compared with their environmental habitat. This suggests that although pre- and post-metamorphic toad life stages differ from each other, the habitat and host fungal communities are so similar that identifying obligate host symbionts is difficult with the techniques used here. This approach does, however, leverage sequence data from host and habitat samples to predict which microbial taxa are host-associated versus transient microbes, thereby condensing a large set of sequence data into a smaller list of potential targets for further consideration.