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  1. Abstract

    Fungal endophytes are pivotal components of a plant's microbiome, profoundly impacting its health and fitness. Yet, myriad questions remain concerning the intricate interactions between these microorganisms and their hosts, particularly in the context of agriculturally important plants such asCoffea arabica. To bridge this knowledge gap and provide a comprehensive framework, this study investigated how farming practices shape the taxonomic and functional diversity of phylloplane endophytes in coffee. Coffee plant leaves from two distinct producing regions in Costa Rica were sampled, ensuring the representation of various coffee varieties (Obatá, Catuaí, and Caturra), agricultural management methods (organic vs. conventional), sun exposure regimes (full sunlight/monoculture vs. natural shade/agroforestry), and leaf developmental stages (newly emerged asymptomatic vs. mature leaves). Fungal communities were characterized by employing both culture‐dependent and independent techniques (internal transcribed spacer 2 nuclear ribosomal DNA metabarcoding). The results showed a greater diversity of endophytes in mature leaves and conventionally managed plants, with coffee variety exerting an unclear influence. The effect of sun exposure was surprisingly negligible. However, data emphasize the benefits of agroforestry and organic farming, which are linked to reduced putative pathogens and heightened levels of potentially mutualistic fungi, fostering functionally diverse communities. Despite the role that plant microbiomes might play in agricultural production, the knowledge to shape endophytic communities through breeding or management is lacking. The results from this study provide a framework to understand how both plant and agricultural practices influence endophyte diversity within coffee crops. These insights hold promise for guiding future efforts to manipulate coffee microbial communities effectively.

     
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    Free, publicly-accessible full text available March 1, 2025
  2. Abstract Aims

    The American leaf spot, caused by Mycena citricolor, is an important disease of coffee (Coffea arabica), mostly in Central America. Currently, there are limited pathogen control alternatives that are environment friendly and economically accessible. The use of fungi isolated from the plant endomycobiota in their native habitats is on the rise because studies show their great potential for biological control. To begin to generate a green alternative to control M. citricolor, the objectives of the present study were to (i) collect, identify, screen (in vitro and in planta), and select endophytic fungi from wild Rubiaceae collected in old-growth forests of Costa Rica; (ii) confirm endophytic colonization in coffee plantlets; (iii) evaluate the effects of the endophytes on plantlet development; and (iv) corroborate the antagonistic ability in planta.

    Methods and results

    Through in vitro and in planta antagonism assays, we found that out of the selected isolates (i.e. Daldinia eschscholzii GU11N, Nectria pseudotrichia GUHN1, Purpureocillium aff. lilacinum CT24, Sarocladium aff. kiliense CT25, Trichoderma rifaii CT5, T. aff. crassum G1C, T. aff. atroviride G7T, T. aff. strigosellum GU12, and Xylaria multiplex GU14T), Trichoderma spp. produced the highest growth inhibition percentages in vitro. Trichoderma isolates CT5 and G1C were then tested in planta using Coffea arabica cv. caturra plantlets. Endophytic colonization was verified, followed by in planta growth promotion and antagonism assays.

    Conclusions

    Results show that Trichoderma isolates CT5 and G1C have potential for plant growth promotion and antagonism against Mycena citricolor, reducing incidence and severity, and preventing plant mortality.

     
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  3. Abstract

    microbeMASST, a taxonomically informed mass spectrometry (MS) search tool, tackles limited microbial metabolite annotation in untargeted metabolomics experiments. Leveraging a curated database of >60,000 microbial monocultures, users can search known and unknown MS/MS spectra and link them to their respective microbial producers via MS/MS fragmentation patterns. Identification of microbe-derived metabolites and relative producers without a priori knowledge will vastly enhance the understanding of microorganisms’ role in ecology and human health.

     
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    Free, publicly-accessible full text available February 5, 2025
  4. Wilson, Richard A. (Ed.)

    Trichodermais a cosmopolitan genus with diverse lifestyles and nutritional modes, including mycotrophy, saprophytism, and endophytism. Previous research has reported greater metabolic gene repertoires in endophytic fungal species compared to closely-related non-endophytes. However, the extent of this ecological trend and its underlying mechanisms are unclear. Some endophytic fungi may also be mycotrophs and have one or more mycoparasitism mechanisms. Mycotrophic endophytes are prominent in certain genera likeTrichoderma, therefore, the mechanisms that enable these fungi to colonize both living plants and fungi may be the result of expanded metabolic gene repertoires. Our objective was to determine what, if any, genomic features are overrepresented in endophytic fungi genomes in order to undercover the genomic underpinning of the fungal endophytic lifestyle. Here we compared metabolic gene cluster and mycoparasitism gene diversity across a dataset of thirty-eightTrichodermagenomes representing the full breadth of environmentalTrichoderma’s diverse lifestyles and nutritional modes. We generated four newTrichoderma endophyticumgenomes to improve the sampling of endophytic isolates from this genus. As predicted, endophyticTrichodermagenomes contained, on average, more total biosynthetic and degradative gene clusters than non-endophytic isolates, suggesting that the ability to create/modify a diversity of metabolites potential is beneficial or necessary to the endophytic fungi. Still, once the phylogenetic signal was taken in consideration, no particular class of metabolic gene cluster was independently associated with theTrichodermaendophytic lifestyle. Several mycoparasitism genes, but no chitinase genes, were associated with endophyticTrichodermagenomes. Most genomic differences betweenTrichodermalifestyles and nutritional modes are difficult to disentangle from phylogenetic divergences among species, suggesting thatTrichodermagenomes maybe particularly well-equipped for lifestyle plasticity. We also consider the role of endophytism in diversifying secondary metabolism after identifying the horizontal transfer of the ergot alkaloid gene cluster toTrichoderma.

     
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    Free, publicly-accessible full text available December 21, 2024
  5. Introduction Products of plant secondary metabolism, such as phenolic compounds, flavonoids, alkaloids, and hormones, play an important role in plant growth, development, stress resistance. The plant family Rubiaceae is extremely diverse and abundant in Central America and contains several economically important genera, e.g. Coffea and other medicinal plants. These are known for the production of bioactive polyphenols (e.g. caffeine and quinine), which have had major impacts on human society. The overall goal of this study was to develop a high-throughput workflow to identify and quantify plant polyphenols. Methods First, a method was optimized to extract over 40 families of phytochemicals. Then, a high-throughput metabolomic platform has been developed to identify and quantify 184 polyphenols in 15 min. Results The current metabolomics study of secondary metabolites was conducted on leaves from one commercial coffee variety and two wild species that also belong to the Rubiaceae family. Global profiling was performed using liquid chromatography high-resolution time-of-flight mass spectrometry. Features whose abundance was significantly different between coffee species were discriminated using statistical analysis and annotated using spectral databases. The identified features were validated by commercially available standards using our newly developed liquid chromatography tandem mass spectrometry method. Discussion Caffeine, trigonelline and theobromine were highly abundant in coffee leaves, as expected. Interestingly, wild Rubiaceae leaves had a higher diversity of phytochemicals in comparison to commercial coffee: defense-related molecules, such as phenylpropanoids (e.g., cinnamic acid), the terpenoid gibberellic acid, and the monolignol sinapaldehyde were found more abundantly in wild Rubiaceae leaves. 
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  6. The transition from conventional to organic agriculture is often challenged by the adaptation of biological control agents to environments heavily exposed to agrochemical pollutants. We studied Trichoderma species isolated from living leaf tissues of wild Rubiacaeae (coffee family) plants to determine their fungicide tolerance and potential for bioremoval. First, we assessed the in vitro tolerance to fungicides of four Trichoderma isolates ( Trichoderma rifaii T1, T . aff. crassum T2, T . aff. atroviride T3, and T . aff. strigosellum T4) by placing mycelial plugs onto solid media supplemented with seven different systemic and non-systemic fungicides. After a week, most of the fungicides did not significantly inhibit the growth of the isolates, except in the case of cyproconazole, where the only isolate able to grow was T1; however, the colony morphology was affected by the presence of fungicides. Second, biological removal potential was established for selected isolates. For this experiment, the isolates T1, T2, and T4 were independently inoculated into liquid media with the fungicides azoxystrobin, chlorothalonil, cyproconazole, and trifloxystrobin. After 14 days of incubation, a removal of up to 89% was achieved for chlorothalonil, 46.4% for cyproconazole, and 33.1% for trifloxystrobin using viable biomass. In the case of azoxystrobin, the highest removal (82.2%) occurred by adsorption to fungal biomass. Ecotoxicological tests in Daphnia magna revealed that T1 has the highest removal potential, achieving significant elimination of every fungicide, while simultaneously detoxifying the aqueous matrix (except in the case of cyproconazole). Isolate T4 also exhibited an intermediate efficiency, while isolate T2 was unable to detoxify the matrix in most cases. The removal and detoxification of cyproconazole failed with all the isolates. These findings suggest that endosphere of wild plants could be an attractive guild to find new Trichoderma species with promising bioremediation capabilities. In addition, the results demonstrate that attention should be placed when combining certain types of agrochemicals with antagonistic fungi in Integrated Pest and Disease Management strategies or when transitioning to organic agriculture. 
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  7. Los estudios ecológicos de las comunidades fúngicas se han visto favorecidos gracias a la aparición y mejora de técnicas independientes de cultivo que utilizan la región ITS como marcador molecular. Esto ha permitido una identificación más precisa en comparación con los métodos tradicionales dependientes de la cultura. Las técnicas de secuenciación de próxima generación han aumentado la cantidad de datos disponibles para la comprensión de las comunidades de hongos endofíticos. Una parte importante de este proceso es la inferencia filogenética para descifrar cómo se relacionan e interactúan los diferentes taxones, sin embargo, este puede convertirse en uno de los análisis bioinformáticos que exige más tiempo. En respuesta a esto, la bioinformática junto con la informática de alto rendimiento ofrecen soluciones para acelerar y hacer más eficientes las herramientas disponibles para el procesamiento de datos a través de la implementación de supercomputadoras y la paralelización de herramientas. En este estudio llevamos a cabo el procesamiento de secuencias ITS para luego utilizar la paralelización de Exabayes, software especializado en el análisis y creación de árboles filogenéticos. Gracias al uso de esta técnica, fue posible reducir el tiempo de ejecución de Exabayes de más de 400 horas a 6 horas, lo que demuestra los beneficios del uso de plataformas informáticas de alto rendimiento. 
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