Endophytic Fungi as a Source of Antiviral Compounds - A Review.

Endophytic fungi are a rich source of secondary metabolites. The interactions between endophytes and their hosts lead to the production of several bioactive substances grouped into different classes, each having a wide variety of effects against various pathogens. The metabolites obtained from these organisms include steroids, alkaloids, phenols, isocoumarins, xanthones, quinones, and terpenoids, among others. These substances are known to have antibiotic, antiparasitic, antifungal, and antiviral effects. This review summarizes secondary metabolites with antiviral effects produced by endophytic fungi and highlights the importance of research in developing novel antiviral substances. We demonstrate that endophytic fungi are a rich source of secondary metabolites that combat pathologies caused by viruses. Optimizing practical and biotechnological screening tools for the research of these metabolites will provide promising drugs to combat these infections.

Agrobacterium-Mediated Transformation of Diaporthe schini Endophytes Associated with Vitis labrusca L. and Its Antagonistic Activity Against Grapevine Phytopathogens.

Fungus-caused diseases are among the greatest losses in grapevine culture. Biological control of pathogens by endophytes may be used to decrease fungicide application rates and environmental impacts. Previously, Diaporthe sp. B46-64 and C27-07 were highlighted as antagonists of grapevine phytopathogens. Herein, molecular multigene (ITS-TUB-TEF1) identification and phylogenetic analysis allowed the identification of these endophytes as belonging to Diaporthe schini species. Agrobacterium tumefaciens-mediated transformation was employed for obtaining 14 stable and traceable gfp- or DsRed-expressing transformants, with high transformation efficiency: 96% for the pFAT-GFP plasmid and 98% for pCAM-DsRed plasmid. Transformants were resistant to hygromycin B with gene hph confirmed by polymerase chain reaction and proved to be mitotically stable, expressing the fluorescent phenotype, with morphological differences in the colonies when compared with wild strains. In vitro antagonism tests revealed an increased antagonistic activity of some transformant strains. The current genetic transformation of D. schini mediated by A. tumefaciens proved to be an efficient technique within the randomized insertion of reporter genes for the monitoring of the strain in the environment.

Bioprospection of Culturable Endophytic Fungi Associated with the Ornamental Plant Pachystachys lutea.

Endophytes are fungi and bacteria that inhabit plant tissues without causing disease. Endophytes have characteristics that are important for the health of the plant and have been isolated from several plants of economic and medicinal interest but rarely from ornamental plants. The current study isolates and identifies endophytic fungi from the leaves of Pachystachys lutea and evaluates the antagonistic activity of these endophytes as well as cellulase production by the endophytes. Fungi were isolated by fragmentation from surface-disinfected leaves and were identified by the sequencing of the ITS gene and the genes coding for EF 1-α and ß-tubulin followed by multilocus sequence analysis. Molecular taxonomic analysis revealed that 78% of the identified fungi belonged to the genus Diaporthe. We also identified strains belonging to the genera Colletotrichum, Phyllosticta, Xylaria, Nemania, and Alternaria. Most of the strains tested were able to inhibit the growth of pathogenic fungi, especially PL09 (Diaporthe sp.), which inhibited the growth of Colletotrichum sp., and PL03 (Diaporthe sp.), which inhibited the growth of Fusarium oxysporum. The production of cellulase ranged from 0.87 to 1.60 µmol/min. Foliar endophytic fungal isolates from P. lutea showed promising results for the in vitro control of plant pathogens and for cellulase production. This paper is the first report on culturable endophytic fungi isolated from the ornamental plant P. lutea.

Endophytic bacterial microbiome associated with leaves of genetically modified (AtAREB1) and conventional (BR 16) soybean plants.

Plant leaves (phyllosphere) have a great potential for colonization and microbial growth, consisting of a dynamic environment in which several factors can interfere with the microbial population structure. The use of genetically modified (GM) plants has introduced several traits in agriculture, such as the improvement of plant drought tolerance, as observed in the AtAREB1 transcription factor overexpression in soybean (Glycine max L. Merrill). The present study aimed at investigating the taxonomic and functional profile of the leaf microbial community of bacteria found in GM (drought-tolerant event 1Ea2939) and conventional (BR 16) soybean plants. Bacterial DNA was extracted from leaf samples collected from each genotype and used for microbial diversity and richness analysis through the MiSeq Illumina platform. Functional prediction was performed using the PICRUSt tool and the STAMP v 2.1.3 software. The obtainment of the GM event 1Ea2939 showed minimum effects on the microbial community and in the potential for chemical-genetic communication, i.e. in the potential for symbiotic and/or mutualistic interaction between plants and their natural microbiota.

Activity of the endophytic fungi Phlebia sp. and Paecilomyces formosus in decolourisation and the reduction of reactive dyes' cytotoxicity in fish erythrocytes.

The current study investigates the potential for discolouration and degradation of Reactive Blue 19 and Reactive Black 5 textile dyes by endophytic fungi Phlebia sp. and Paecilomyces formosus as well as the potential cytotoxicity of products or by-products generated by the treatments in fish erythrocytes. It was observed at 30 days that both endophytes showed biodegradation activity with 0.1 g mL-1 of dyes. P. formosus showed highest extracellular and intracellular protein content levels after the 15th day, and Phlebia sp. stands out for production of extracellular laccase, indicating that this enzyme may be associated with the decolouration capacity. The dyes showed toxic effects in fishes at 0.01 g mL-1 concentration, resulting in the appearance of micronuclei in erythrocyte cells. When degraded dyes treated by endophytes were tested, the frequency of micronuclei reduced approximately 20%, indicating the effectiveness of these endophytic in the treatment of textile dyes with less environmental impact, thus indicating a potential for application of these fungi in bioremediation process.

Endophytic cultivable bacterial community obtained from the Paullinia cupana seed in Amazonas and Bahia regions and its antagonistic effects against Colletotrichum gloeosporioides.

Guarana (Paullinia cupana var. sorbilis) is a plant from the Amazonas region with socio-economic importance. However, guarana production has been increasingly affected by unfavorable conditions resulting from anthracnose, caused by the Colletotrichum fungal genus, which primarily affects mainly the Amazonas region. The aim of the present study was to isolate bacterial endophytes from the seeds of guarana plants obtained from Amazonas region and the Northeast state of Bahia, a region where this disease is not a problem for guarana plantations. The number of bacterial Colony Forming Units (CFU/g seeds) was 2.4 × 10(4) from the Bahia and 2.9 × 10(4) from the Amazonas region. One hundred and two isolated bacteria were evaluated in vitro against the phytopathogenic strain Colletotrichum gloeosporioides L1. These isolates were also analyzed for the enzymatic production of amylase, cellulase, protease, pectinase, lipase and esterase. Approximately 15% of isolates, showing high antagonistic activity, and the production of at least one enzyme were identified through the partial sequencing of 16S rDNA. The genus Bacillus was the most frequently observed, followed by Paenibacillus, Ochrobactrum, Microbacterium and Stenotrophomonas. Proteolytic activity was observed in 24 isolates followed by amylolytic, pectinolytic and cellulolytic activities. No esterase and lipase production was detected. Most of the isolates, showing antagonistic effects against C. gloeosporioides and high enzymatic activities, were isolated from the anthracnose-affected region. A biocontrol method using the endophytes from guarana seeds could be applied in the future, as these bacteria are vertically transferred to guarana seedlings.

Radiotolerant endophytic bacteria and analysis of the effects of 137Cesium on the metabolome of Pantoea sp.

Some bacteria have developed mechanisms to withstand the stress caused by ionizing radiation. The ability of these radioresistant microorganisms to survive high levels of radiation is primarily attributed to their DNA repair mechanisms and the production of protective metabolites. To determine the effect of irradiation on bacterial growth, we propose to compare the metabolites produced by the irradiated isolates to those of the control (non-irradiated isolates) using mass spectrometry, molecular networking, and chemometric analysis. We identified the secondary metabolites produced by these bacteria and observed variations in growth following irradiation. Notably, after 48 h of exposure to radiation, Pantoea sp. bacterial cells exhibited a significant 6-log increase compared to non-irradiated cells. Non-irradiated cells produce exclusively Pyridindolol, 1-hydroxy-4-methylcarbostyril, N-alkyl, and N-2-alkoxyethyl diethanolamine, while 5'-methylthioadenosine was detected only in irradiated cells. These findings suggest that the metabolic profile of Pantoea sp. remained relatively stable. The results obtained from this study have the potential to facilitate the development of innovative strategies for harnessing the capabilities of endophytic bacteria in radiological protection and bioremediation of radionuclides.

Synthesis, characterization, and reusability of novel nanobiocomposite of endophytic fungus Aspergillus flavus and magnetic nanoparticles (Fe3O4) with dye bioremediation potential.

The incorrect disposal of textile dyes, such as Reactive Black 5 (RB5), causes several problems for living beings and the quality of the environment. Nanobiocomposites (NBC) produced from endophytic fungi (potentially remediation dyes-agents) and magnetic nanoparticles have high biotechnological potential due to their superparamagnetic behavior, which would allow their recovery through the magnetic field after the bioremediation process. This work aimed to obtain a new nanobiocomposite from the interaction of magnetite nanoparticles (Fe3O4) with the endophyte Aspergillus flavus (Af-CL-7) to evaluate its bioremediation capacity and to reduce the toxicity of RB5 and its reuse. Before obtaining the NBC, Af-CL-7 showed discoloration of RB5 and it was tolerant to all tested concentrations of this dye. The discovery of the nanobiocomposite textile dye bioremediator product presents a significant environmental advantage by addressing the issue of water pollution caused by textile dyes. The NBC called Af-Fe3O4 was successfully obtained with the magnetized endophyte, and their magnetic properties were verified by VSM analysis and by action of magnetic fields generated by Nd-Fe-B magnets SEM analyzes showed that the nanoparticles did not cause any damage to the hypha morphology, and TEM analyzes confirmed the presence of nanoparticles in the fungus wall and also inside the cell. The NBC Af-Fe3O4 and Af-CL-7 showed, respectively, 96.1% and 92.2% of RB5 discoloration in the first use, 91.1% e 86.2% of discoloration in the validation test, and 89.0% in NBC reuse. In the toxicological bioassay with Lactuca sativa seeds, NBC showed a positive reduction in the toxicity of RB5 after treatment, allowing the hypocotyl growth to be statistically similar to the control with water. Thus, we highlight the promising obtaining process of NBC that could be applied in bioremediation of contaminated waters, wherein the industrial economic cost will depend on the fermentation efficiency, biomass production and nanoparticle synthesis.

Antitumor and antibacterial activity of metabolites of endophytic Colletotrichum siamense isolated from coffee (Coffea arabica L. cv IAPAR-59).

Endophytic fungi produce a range of known metabolites and several others, not yet explored, which present important biological activities from the pharmaceutical and industrial perspective. Several studies have reported the diversity of endophytes in Coffea arabica plants, although few have been described in organic cultures. In the current paper, we describe the chemical profile of specialized metabolites in the ethyl acetate phase in a strain of the endophytic fungus Colletotrichum siamense associated with coffee (Coffea arabica L.) (Rubiaceae) and its potential against tumor cells and bacteria of medical and food importance. Cytotoxicity assays in tumor cells MCF-7 and HepG2/C3A were performed by MTT and microdilution in broth to evaluate the antibacterial action of metabolic extract. The antiproliferative assay showed promising results after 24 h of treatment, with 50% injunction concentrations for the two cell types. UHPLC-MS/MS analyses with an electrospray ionization source were used to analyze the extracts and identify compounds of species Colletotrichum siamense, which is still little explored as a source of active metabolites. Many of these compounds observed in the endophytic need to be chemically synthesized in industry, at high costs, while production by the fungus becomes a chemically and economically more viable alternative. Pyrocatechol, gentisyl alcohol, and alpha-linolenic acid, associated with different mechanisms of action against tumor cells, were detected among the main compounds. The extract of the endophytic fungus Colletotrichum siamense presented several compounds with pharmacological potential and antibacterial activity, corroborating its potential in biotechnological applications.

Molecular techniques for the taxonomy of Aedes Meigen, 1818 (Culicidae: Aedini): A review of studies from 2010 to 2021.

The original description of Aedes Meigen in 1818, written in Latin, was very brief and included a single species, Aedes cinereus. In the last two decades the genus Aedes (Meigen, 1818) has undergone several revisions and reclassifications, with the current proposal being described by Wilkerson in 2015. However, the available keys for morphological identification are still not sufficient to differentiate cryptic species, damaged species, or those with confusing taxonomy. The current study aims to identify and describe the main taxonomic proposals and molecular methodologies available for the identification of the genus Aedes published between the years 2010 and 2021. The main molecular techniques used to identify the genus in the last 10 years, are: Multiplex PCR, DNA barcoding, nuclear and mitochondrial markers, environmental DNA, and bacterial microbiome analysis. This review highlights that there are catalogued data for only a few species of the genus Aedes, being restricted to medically important taxa such as Aedes albopictus and Aedes aegypti. The integrative taxonomy approach is a possibility to reconcile morphological and molecular data to improve species delimitation, contributing to future revisions of the genus.

Differential Chemical Profile of Metabolite Extracts Produced by the Diaporthe citri (G-01) Endophyte Mediated by Varying the Fermented Broth pH.

Endophytic microorganisms show great potential for biotechnological exploitation because they are able to produce a wide range of secondary compounds involved in endophyte−plant adaptation, and their interactions with other living organisms that share the same microhabitat. Techniques used to chemically extract these compounds often neglect the intrinsic chemical characteristics of the molecules involved, such as the ability to form conjugate acids or bases and how they influence the solubilities of these molecules in organic solvents. Therefore, in this study, we aimed to evaluate how the pH of the fermented broth affects the process used to extract the secondary metabolites of the Diaporthe citri strain G-01 endophyte with ethyl acetate as the organic solvent. The analyzed samples, conducted by direct-infusion electrospray-ionization mass spectrometry, were grouped according to the pH of the fermented broth (i.e., <7 and ≥7). A more extreme pH (i.e., 2 or 12) was found to affect the chemical profile of the sample. Moreover, statistical analysis enabled us to determine the presence or absence of ions of high importance; for example, ions at 390.7 and 456.5 m/z were observed mainly at acidic pH, while 226.5, 298.3, and 430.1 m/z ions were observed at pH ≥ 7. Extraction at a pH between 4 and 9 may be of interest for exploring the differential secondary metabolites produced by endophytes. Furthermore, pH influences the chemical phenotype of the fungal metabolic extract.

Influence of plant growth-promoting endophytes Colletotrichum siamense and Diaporthe masirevici on tomato plants (Lycopersicon esculentum Mill.).

The protective and growth-promoting activities of Colletrotrichum and Diaporthe endophytes on tomato plants (Lycopersicon esculentum Mill.) are underexplored. We screened 40 endophytic fungi associated with Mexican shrimp plant (Justicia brandegeana) using an in vitro dual culture assay for Fusarium oxysporum, one of the most important phytopathogens of tomato plants. The three best antagonists, Colletotrichum siamense (JB224.g1), C. siamense (JB252.g1), and Diaporthe masirevicii (JB270), were identified based on multilocus sequence analysis. They were assessed in vitro for their inhibition of F. oxysporum and phosphate solubilisation capacity, and for the production of indole acetic acid. Greenhouse experiments verified the growth-promoting effects of these endophytes and the suppression of F. oxysporum symptoms in tomato plants.   Under greenhouse conditions, the JB252.g1 and JB270 isolates showed positive results for seedling emergence speed. The radicular system depth of plants inoculated with JB270 was greater than that in uninoculated plants (27.21 vs 21.95 cm). The soil plant analysis development chlorophyll metre (SPAD) index showed statistically significant results, especially for the endophyte JB224.g1 (36.99) compared to the control plants (30.90) and plants infected solely with F. oxysporum (33.64).

Antimicrobial and antioxidant activities of secondary metabolites from endophytic fungus Botryosphaeria fabicerciana (MGN23-3) associated to Morus nigra L.

This study was to evaluate the biological activity of the extract of Botryosphaeria fabicerciana isolated from leaves of Morus nigra. The volatile compounds from the crude extract were analysed by GC-MS which demonstrate that mellein and ß-orcinaldehyde were are the major compounds. The best minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the extract was observed against Gram-positive bacteria, with a MIC of 15.6 µg/mL towards B. cereus and MIC of 62.5 µg/mL towards S. aureus and B. subtilis. MBC values of 31.25 µg/mL, 62.5 µg/mL, and 250 µg/mL were observed towards B. cereus, B. subtilis, and S. aureus, respectively. The cytotoxicity analyses showed CC50 of 115 µg/mL. The crude extract showed antioxidant activity by the DPPH, ABTS, and FRAP assays. Therefore, the extract of the endophytic fungus presented biotechnological potential as an antibacterial and antioxidant agent.

Retrotransposons and multilocus sequence analysis reveals diversity and genetic variability in endophytic fungi-associated with Serjania laruotteana Cambess.

The composition of endophytic communities is dynamic and demonstrates host specificity; besides, they have great intra- and interspecific genetic variability. In this work, we isolated leaf endophytic fungi from Serjania laruotteana, identify them using multilocus analysis, and evaluate the genetic variability using IRAP (inter-retrotransposon amplified polymorphism) and REMAP (retrotransposon-microssatellite amplified polymorphism). A total of 261 fungi were isolated and 58 were identified. Multilocus phylogenetic analysis using the partial sequences from the ITS1-5.8S-ITS2 regions, elongation factor 1-alpha, ß-tubulin, actin, glyceraldehyde-3-phosphate dehydrogenase, and calmodulin genes identify that most strains belonged to the Colletotrichum and Diaporthe genera, other isolated genera were Xylaria, Phyllosticta, Muyocopron, Fusarium, Nemania, Plectosphaerella, Corynespora, Bipolaris, and Curvularia. The IRAP and REMAP analyzes were performed with Colletotrichum and Diaporthe genera and showed 100% of polymorphism and high intra- and interspecific variability. This is the first report of the diversity of endophytic fungi from S. laruotteana. In addition, it demonstrated that the IRAP and REMAP can be used to distinguish morphologically similar lineages, revealing differences even strains of the same species.

Secondary metabolites of Curvularia sp. G6-32, an endophyte of Sapindus saponaria, with antioxidant and anticholinesterasic properties.

In the present study, the biological activity of an extract of the secondary metabolites (E-G6-32) produced by the Curvularia sp. G6-32 endophyte (isolated from the medicinal plant Sapindus saponaria L.) was investigated. The antioxidant potential was confirmed by the DPPH (22.5%) and ABTS (62.7%) assays, and the total phenolic compound content was 40 µg gallic acid equivalents/mg. The extract E-G6-32 displayed good inhibitory activity toward butyrylcholinesterase (BuChE; IC50 = 110 ± 0.05 µg mL-1). The extract E-G6-32 was subjected to spectroscopic and mass spectrometry analyses. Comparison with the literature data confirmed that (-)-asperpentyn (1) was a major component. Asperpentyn belongs to the epoxyquinone family, which has attractive structural complexity, diverse functional groups, and a broad range of biological activities, including specific enzyme inhibitory activity. Our results suggest that Curvularia sp. G6-32 is a promising source of bioactive secondary metabolites and contains (-)-asperpentyn, which has potential pharmaceutical interest.[Figure: see text].

Metabolic extract of the endophytic fungus Flavodon flavus isolated from Justicia brandegeana in the control of Alicyclobacillus acidoterrestris in commercial orange juice.

In this work, the antibacterial activity of a crude extract of the endophytic fungus Flavodon flavus (JB257), isolated from leaves of Justicia brandegeana, was evaluated against both the vegetative and sporulated forms of Alicyclobacillus acidoterrestris. The microdilution technique was performed in order to determine the antibacterial activity of the crude extract alone as well as in combination with the bacteriocin, nisin. The minimum inhibitory concentration (MIC) of the crude extract and nisin alone against A. acidoterrestris vegetative forms were 250 µg/mL and 31.5 µg/mL, respectively, while the minimum bactericidal concentrations (MBC) were 1000 µg/mL and 62.5 µg/mL,respectively. For A. acidoterrestris spores, treatment with the crude extract at a concentration of 500 µg/mL caused a 47% reduction in growth, while nisin at 62.5 µg/mL could reduce 100% of the growth. The in vitro evaluation of the crude extract combined with nisin against A. acidoterrestris by the Checkerboard method showed a synergistic interaction between the two compounds. In addition, greater selectivity towards bacterial cells over host cells, a human hepatocyte cell line, was achieved when the crude extract was combined with nisin, Using scanning electron microscopy, interferences in the cell membrane of A. acidoterrestris could be observed after treatment with the crude extract. The results presented in this study indicate that the crude extract of the endophyte F. flavus has biotechnological potential in the food industry, especially for the treatment of orange juices through the control of A. acidoterrestris.

Tibouchina granulosa (Vell.) Cogn (Melastomataceae) as source of endophytic fungi: isolation, identification, and antiprotozoal activity of metabolites from Phyllosticta capitalensis.

Endophytes are microorganisms that form symbiotic relationships with their own host. Included in this group are the species Phyllosticta capitalensis, a group of fungi that include saprobes that produce bioactive metabolites. The present study aimed to identify the cultivable endophytic fungal microbiota present in healthy leaves of Tibouchina granulosa (Desr.) Cogn. (Melastomataceae) and investigate secondary metabolites produced by a strain of P. capitalensis and their effects against both Leishmania species and Trypanossoma cruzi. Identification of the strains was accomplished through multilocus sequencing analysis (MLSA), followed by phylogenetic analysis. The frequency of colonization was 73.66% and identified fungi belonged to the genus Diaporthe, Colletotrichum, Phyllosticta, Xylaria, Hypoxylon, Fusarium, Nigrospora, and Cercospora. A total of 18 compounds were identified by high-resolution mass spectrum analysis (UHPLC-HRMS), including fatty acids based on linoleic acid and derivatives, from P. capitalensis. Crude extracts had activity against Leishmania amazonensis, L. infantum, and Trypanosoma cruzi, with inhibitory concentration (IC50) values of 17.2 µg/mL, 82.0 µg/mL, and 50.13 µg/mL, respectively. This is the first report of the production of these compounds by the endophytic P. capitalensis isolated from T. granulosa.

Curvulin and spirostaphylotrichins R and U from extracts produced by two endophytic Bipolaris sp. associated to aquatic macrophytes with antileishmanial activity.

In the present study, biological activity and chemical composition of two crude extracts of endophytic fungal strains of Bipolaris genera isolated from two species of aquatic macrophytes: Eichhornia azurea (Kunth) and Eichhornia crassipes (Mart.) were investigated. The nuclear magnetic resonance and mass spectrometry data provided the identification of three main compounds: curvulin (1), spirostaphylotrichin R (2) and U (3). The fragmentation mechanism of the precursor ions towards collision induced dissociation (CID) tandem mass spectrometry experiment (MS/MS) is also proposed. Furthermore, biological screening of the crude extracts displayed antileishmanial activity with IC50 values ranging from 70-84.2 µg.mL-1.

3-Nitropropionic acid production by the endophytic Diaporthe citri: Molecular taxonomy, chemical characterization, and quantification under pH variation.

3-nitropropionic acid (3-NPA) is a nitrogenated compound produced by plants and fungi and has been associated with poisoning episodes in humans, animals, and to induction of Huntington disease symptoms in rats. The production of 3-NPA by endophytes has been reported, but the function and biosynthesis are not well-defined. The specie of endophytic strain G-01 was confirmed as Diaporthe citri using a multilocus sequence analysis, and was verified different concentrations of 3-NPA produced at different initial pHs by these strain. The chemical analysis indicated that 3-NPA was the majority compound present in the crude extracts. The better extraction condition was at an initial pH of 7.0 for 22 d, yielding about 80 % of 3-NPA per mg of extract. It was observed that the concentration of 3-NPA increased after the initial consumption of reduction sugars, indicating that the compound is produced after the high energetic production phase of the fungus. These and other studies demonstrate the production of this compound by plants and endophytic fungi, indicating that 3-NPA may be involved in defence and nutrition systems of endophytes and host plants, and they also might participate in the biogeochemical nitrogen cycle.

Antifungal and proteolytic activities of endophytic fungi isolated from Piper hispidum Sw.

Endophytes are being considered for use in biological control, and the enzymes they secrete might facilitate their initial colonization of internal plant tissues and direct interactions with microbial pathogens. Microbial proteases are also biotechnologically important products employed in bioremediation processes, cosmetics, and the pharmaceutical, photographic and food industries. In the present study, we evaluated antagonism and competitive interactions between 98 fungal endophytes and Alternaria alternata, Colletotrichum sp., Phyllosticta citricarpa and Moniliophthora perniciosa. We also examined the proteolytic activities of endophytes grown in liquid medium and conducted cup plate assays. The results showed that certain strains in the assemblage of P. hispidum endophytes are important sources of antifungal properties, primarily Lasiodiplodia theobromae JF766989, which reduced phytopathogen growth by approximately 54 to 65%. We detected 28 endophytes producing enzymatic halos of up to 16.40 mm in diameter. The results obtained in the present study highlight the proteolytic activity of the endophytes Phoma herbarum JF766995 and Schizophyllum commune JF766994, which presented the highest enzymatic halo diameters under at least one culture condition tested. The increased activities of certain isolates in the presence of rice or soy flour as a substrate (with halos up to 17.67 mm in diameter) suggests that these endophytes have the potential to produce enzymes using agricultural wastes.