Identification of Secondary Metabolites from the Mangrove-Endophyte Lasiodiplodia iranensis F0619 by UPLC-ESI-MS/MS.

Lasiodiplodia is a widely distributed fungal genus, frequently found in tropical and subtropical regions where it can cause disease in important crops. It represents a promising source of active secondary metabolites with uses in chemical, pharmaceutical, and agrochemical processes. In this study, the strain Lasiodiplodia iranensis F0619 was isolated from the mangrove Avicennia ger-minans, collected from Sarigua National Park in the Republic of Panama. Fractions of crude extract were analyzed by UPLC-ESI-MS/MS, and five compounds, previously reported from Lasiodiplodia genus were identified, including 11,12-didehydro-7-iso-jasmonic acid (1), 4,5-didehydro-7-iso-jasmonic acid (2), cyclo-(L-Leu-L-Pro) (3), jasmonate-threonine (4), and abscisic acid (5). We describe and analyze their MS/MS fragmentation patterns to confirm the compounds 'chemical structures.

Structurally Uncommon Secondary Metabolites Derived from Endophytic Fungi.

Among microorganisms, endophytic fungi are the least studied, but they have attracted attention due to their high biological diversity and ability to produce novel and bioactive secondary metabolites to protect their host plant against biotic and abiotic stress. These compounds belong to different structural classes, such as alkaloids, peptides, terpenoids, polyketides, and steroids, which could present significant biological activities that are useful for pharmacological or medical applications. Recent reviews on endophytic fungi have mainly focused on the production of novel bioactive compounds. Here, we focus on compounds produced by endophytic fungi, reported with uncommon bioactive structures, establishing the neighbor net and diversity of endophytic fungi. The review includes compounds published from January 2015 to December 2020 that were catalogued as unprecedented, rare, uncommon, or possessing novel structural skeletons from more than 39 different genera, with Aspergillus and Penicillium being the most mentioned. They were reported as displaying cytotoxic, antitumor, antimicrobial, antiviral, or anti-inflammatory activity. The solid culture, using rice as a carbon source, was the most common medium utilized in the fermentation process when this type of compound was isolated.

Antileishmanial macrolides from ant-associated Streptomyces sp. ISID311.

Three antifungal macrolides cyphomycin (1), caniferolide C (2) and GT-35 (3) were isolated from Streptomyces sp. ISID311, a bacterial symbiont associated with Cyphomyrmex fungus-growing ants. The planar structures of these compounds were established by 1 and 2D NMR data and MS analysis. The relative configurations of 1-3 were established using Kishi's universal NMR database method, NOE/ROE analysis and coupling constants analysis assisted by comparisons with NMR data of related compounds. Detailed bioinformatic analysis of cyphomycin biosynthetic gene cluster confirmed the stereochemical assignments. Compounds 1-3 displayed high antagonism against different strains of Escovopsis sp., pathogen fungi specialized to the fungus-growing ant system. Compounds 1-3 also exhibited potent antiprotozoal activity against intracellular amastigotes of the human parasite Leishmania donovani with IC50 values of 2.32, 0.091 and 0.073 µM, respectively, with high selectivity indexes.

Patents on Endophytic Fungi for Agriculture and Bio- and Phytoremediation Applications.

Plant endophytic fungi spend all or part of their lives inside host tissues without causing disease symptoms. They can colonize the plant to protect against predators, pathogens and abiotic stresses generated by drought, salinity, high concentrations of heavy metals, UV radiation and temperature fluctuations. They can also promote plant growth through the biosynthesis of phytohormones and nutrient acquisition. In recent years, the study of endophytic fungi for biological control of plant diseases and pests has been intensified to try to reduce the ecological and public health impacts due the use of chemicals and the emergence of fungicide resistance. In this review, we examine 185 patents related to endophytic fungi (from January 1988 to December 2019) and discuss their applicability for abiotic stress tolerance and growth promotion of plants, as agents for biocontrol of herbivores and plant pathogens and bio- and phytoremediation applications.

Patents on Endophytic Fungi Related to Secondary Metabolites and Biotransformation Applications.

Endophytic fungi are an important group of microorganisms and one of the least studied. They enhance their host's resistance against abiotic stress, disease, insects, pathogens and mammalian herbivores by producing secondary metabolites with a wide spectrum of biological activity. Therefore, they could be an alternative source of secondary metabolites for applications in medicine, pharmacy and agriculture. In this review, we analyzed patents related to the production of secondary metabolites and biotransformation processes through endophytic fungi and their fields of application. We examined 245 patents (224 related to secondary metabolite production and 21 for biotransformation). The most patented fungi in the development of these applications belong to the Aspergillus, Fusarium, Trichoderma, Penicillium, and Phomopsis genera and cover uses in the biomedicine, agriculture, food, and biotechnology industries.

Antifungal compounds from Streptomyces associated with attine ants also inhibit Leishmania donovani.

Bacterial strains isolated from attine ants showed activity against the insect specialized fungal pathogen Escovopsis and also against the human protozoan parasite Leishmania donovani. The bioassay guided fractionation of extracts from cultures of Streptomyces sp. ICBG292, isolated from the exoskeleton of Cyphomyrmex workers, led to the isolation of Mer-A2026B (1), piericidin-A1 (2) and nigericin (3). Nigericin (3) presented high activity against intracellular amastigotes of L. donovani (IC50 0.129 ± 0.008 µM). Streptomyces puniceus ICBG378, isolated from workers of Acromyrmex rugosus rugosus, produced dinactin (4) with potent anti-L. donovani activity against intracellular amastigotes (IC50 0.018 ± 0.003 µM). Compounds 3 and 4 showed good selectivity indexes, 88.91 and 656.11 respectively, and were more active than positive control, miltefosine. Compounds 1-4 were also active against some Escovopsis strains. Compounds 1 and 2 were also produced by Streptomyces sp. ICBG233, isolated from workers of Atta sexdens, and detected in ants' extracts by mass spectrometry, suggesting they are produced in the natural environment as defensive compounds involved in the symbiotic interaction.

The antimicrobial potential of Streptomyces from insect microbiomes.

Antimicrobial resistance is a global health crisis and few novel antimicrobials have been discovered in recent decades. Natural products, particularly from Streptomyces, are the source of most antimicrobials, yet discovery campaigns focusing on Streptomyces from the soil largely rediscover known compounds. Investigation of understudied and symbiotic sources has seen some success, yet no studies have systematically explored microbiomes for antimicrobials. Here we assess the distinct evolutionary lineages of Streptomyces from insect microbiomes as a source of new antimicrobials through large-scale isolations, bioactivity assays, genomics, metabolomics, and in vivo infection models. Insect-associated Streptomyces inhibit antimicrobial-resistant pathogens more than soil Streptomyces. Genomics and metabolomics reveal their diverse biosynthetic capabilities. Further, we describe cyphomycin, a new molecule active against multidrug resistant fungal pathogens. The evolutionary trajectories of Streptomyces from the insect microbiome influence their biosynthetic potential and ability to inhibit resistant pathogens, supporting the promise of this source in augmenting future antimicrobial discovery.

Absolute Configurations of Griseorhodins A and C.

The known antibiotic and cytotoxic compounds griseorhodin A (1) and griseorhodin C (2) were produced in solid culture by Streptomyces puniceus AB10, which was isolated from the leaf-cutter ant Acromyrmex rugosus rugosus. Their absolute configurations were unambiguously established as 6S,6aR,7S,8S and 6R,6aR,7S,8R, respectively, using vibrational circular dichroism (VCD) and density functional theory (DFT) calculations.

Anti-L. donovani activity in macrophage/amastigote model of palmarumycin CP18 and its large scale production.

Palmarumycin CP18, isolated from an extract of the fermentation broth and mycelium of the Panamanian endophytic fungus Edenia sp., was previously reported with strong and specific activity against Leishmania donovani. Here we report that when the same strain was cultured on different solid media--Harrold Agar, Leonian Agar, Potato dextrose Agar (PDA), Corn Meal Agar, Honey Peptone Agar, and eight vegetables (V8) Agar--in order to determine the optimal conditions for isolation of palmarumycin CP18, no signal for this compound was observed in any of the 1H NMR spectra of fractions obtained from these extracts. However, one extract, prepared from the fungal culture in PDA contained significant amounts of CJ-12,372, a possible biosynthetic precursor of palmarumycin CP18. Edenia sp. was cultivated on a large scale on PDA and CJ-12,372 was converted to palmarumycin CP18 by oxidation of its p-hydroquinone moiety with DDQ in dioxane. Palmarumycin CP18 showed anti-leishmanial activity against L. donovani in a macrophage/amastigote model, with IC50 values of 23.5 microM.

Mycoleptodiscins A and B, cytotoxic alkaloids from the endophytic fungus Mycoleptodiscus sp. F0194.

Two novel reddish-orange alkaloids, mycoleptodiscin A (1) and mycoleptodiscin B (2), were isolated from liquid cultures of the endophytic fungus Mycoleptodiscus sp. that had been isolated from Desmotes incomparabilis in Panama. Elucidation of their structures was accomplished using 1D and 2D NMR spectroscopy in combination with IR spectroscopic and MS data. These compounds are indole-terpenes with a new skeleton uncommon in nature. Mycoleptodiscin B (2) was active in inhibiting the growth of cancer cell lines with IC50 values in the range 0.60-0.78 µM.