Biodiversity and application prospects of fungal endophytes in the agarwood-producing genera,Aquilaria and Gyrinops (Thymelaeaceae): A review

Agarwood is originated from the resinous part of Aquilaria and Gyrinops plants and has been a precious biomaterial for applications in traditional medicine, perfumery, cosmetics, and religious purposes all over the world. In the wild, the formation of agarwood is related to the defense mechanism of the tree in response to physical damage that allows further microbial infestation into its wood, while having the whole tree covered with agarwood would take up a long time, and it rarely happens. For Aquilaria and Gyrinops, the presence of endophytes is mainly found derived from the tree. The isolated endophytes could be important sources of natural products, while some could contribute to the formation of agarwood in the tree, which is safe for the environment and human health. This review summarized the biodiversity of fungal endophytes recorded in Aquilaria and Gyrinops and their potential effects on host trees. Till now, 67 endophytic genera have been isolated from Aquilaria and Gyrinops, and 18 ones were found responsible for the promotion of agarwood formation. Additionally, 92 compounds have been reported to be produced by the agarwood endophytes, and 52 ones displayed biological activities, most of which have anti-inflammatory, anti-bacterial, and anti-cancer activities. Nevertheless, fungal endophytes are promising agents that deserved to be further studied and scaled up to a commercial level for the production of agarwood oil, but the role of endophytes in the agarwood host trees needs to be furtherly investigated in future studies.     

Discovery and efficient synthesis of a biologically active alkaloid inspired by thiostrepton biosynthesis

Thiostrepton, a natural peptide macrocycle, is of great interest due to its structural complexity and numerous biological activities, including anti-bacterial, anti-tumor, and anti-plasmodial activities. The quinaldic acid (QA) moiety-containing side ring (loop 2) was proven to play an important role in carrying out these functions. Previously, we proposed biosynthetic logic for thiostrepton loop 2 and demonstrated the formation mechanism of QA. Herein, we report the discovery and efficient synthesis of a biologically active alkaloid, that is, a key intermediate involved in the thiostrepton biosynthetic pathway. A chemo-enzymatic method was performed to synthesize the molecule, and a series of analogs were prepared for bioassays, which included the examination of anti-bacterial and anti-tumor activities.     

Micro-scale extraction procedure for standardized screening of fungal metabolite production in cultures

A simple and rapid standardized micro-scale extraction procedure has been developed to prepare extracts from fungal cultures for high-performance liquid chromatographic (HPLC) analysis. The method is based on ultrasonic extraction of three 6-mm plugs cut from a culture using 0.5 ml of solvent followed by a simple solvent change, filtration and injection. Approximately 5 min of work is involved in the extraction and work-up process and the extract can prepared for HPLC analysis within 60–70 min. The method has been used for determination of chromatographic metabolite profiles from 395 fungal isolates, including all terverticillate Penicillium species, cultivated on both Czapek Yeast Autolysate agar and Yeast Extract Sucrose agar. The concentration of the extracts proved to be sufficient to determine all secondary metabolites reported to be produced by these species using HPLC with diode array detection. These findings were confirmed by analyses of 132 pure metabolite standards.     

Advances in engineering methylotrophic yeast for biosynthesis of valuable chemicals from methanol

Methylotrophic yeasts, which can use methanol as carbon and energy source, have been wildly used as microbial cell factories for biomanufacturing. Methanol derived from diverse sources could be transformed into precursor, such as pyruvate and acetyl-CoA, for the production of valuable chemicals through genetic engineering of methylotrophic yeast.     

From NGS assembly challenges to instability of fungal mitochondrial genomes: A case study in genome complexity

The presence of repetitive or non-unique DNA persisting over sizable regions of a eukaryotic genome can hinder the genome's successful de novo assembly from short reads: ambiguities in assigning genome locations to the non-unique subsequences can result in premature termination of contigs and thus overfragmented assemblies. Fungal mitochondrial (mtDNA) genomes are compact (typically less than 100 kb), yet often contain short non-unique sequences that can be shown to impede their successful de novo assembly in silico. Such repeats can also confuse processes in the cell in vivo. A well-studied example is ectopic (out-of-register, illegitimate) recombination associated with repeat pairs, which can lead to deletion of functionally important genes that are located between the repeats. Repeats that remain conserved over micro- or macroevolutionary timescales despite such risks may indicate functionally or structurally (e.g., for replication) important regions. This principle could form the basis of a mining strategy for accelerating discovery of function in genome sequences. We present here our screening of a sample of 11 fully sequenced fungal mitochondrial genomes by observing where exact k-mer repeats occurred several times; initial analyses motivated us to focus on 17-mers occurring more than three times. Based on the diverse repeats we observe, we propose that such screening may serve as an efficient expedient for gaining a rapid but representative first insight into the repeat landscapes of sparsely characterized mitochondrial chromosomes. Our matching of the flagged repeats to previously reported regions of interest supports the idea that systems of persisting, non-trivial repeats in genomes can often highlight features meriting further attention.     

Effect of chemical compounds on amino acid content of some Fusarium species and its significance to fungal chemotaxonomy

The cellular amino acid profiles of nine species of Fusarium; namely, Fusarium anthophilum, Fusarium avenaceum, Fusarium cerealis, Fusarium graminearum, Fusarium graminum, Fusarium oxysporum f. sp. conglutinans, Fusarium pseudograminearum, Fusarium roseum, and Fusariumsacchari var. elongatum growing on malt extract medium were determined. The amino acid profiles of the investigated fungi were varied and could be used for identification and characterisation of certain Fusarium species. Addition of certain chemical compounds including aspartic acid, glutamic acid, methionine, selenium, and urea to the growth medium affected the amino acid profiles. However, susceptibility of amino acid content to environmental conditions increased the variation of amino acid profiles among all the investigated Fusarium species. Some amino acids were only produced when certain chemical compounds were added to the growth medium. Valine was produced by F. anthophilum only in the presence of aspartic acid or selenium, while serine was produced in the presence of aspartic acid, glutamic acid, or methionine. Also, cysteine was produced by F. avenaceum in the presence of glutamic acid or urea. F. cerealis produced tryptophan only in the presence of aspartic acid or urea, while F. graminearum produced leucine in glutamic, methionine or urea. Similarly, many different amino acids were produced by each Fusarium species only in the presence of certain chemical compounds. The results revealed that the amino acid profiles will be more useful for characterisation and identification of fungi if they are determined under different conditions.     

Aspercyclide A-C, three novel fungal metabolites from Aspergillus sp. as inhibitors of high-affinity IgE receptor

Immunoglobin E (IgE) binds to the high-affinity IgE receptor on mast cells and basofils and causes release of inflammatory compounds that leads to allergic diseases. Inhibition of the ligand binding could lead to blockade of the release of inflammation causing compounds and thus alleviate asthma and other allergic diseases. Natural product screening and bioassay-guided isolation of an extract of Aspergillus sp. led to the identification of three novel 11-membered macrocyclic biphenyl ether lactones, aspercyclides A-C. Aspercyclide A inhibited the IgE binding with an IC₅₀ of 200 μM. The isolation, structure elucidation, absolute stereochemistry, and the binding activities of these compounds are described.     

Ti(III)-induced radical cyclization. A stereoselective entry to the perhydrobenzo[e]indene unit of new protein farnesyltransferase inhibitors, andrastins A-D

An efficient approach for the synthesis of a fully functionalized perhydrobenzo[e]indene, the BCD ring system of andrastins A-D, is described. The synthesis commences from (±)-Wieland-Miescher ketone and features a Ti(III)-induced radical cyclization as the central step.     

Asymmetric Total Synthesis of ent‐Pyripyropene A

An asymmetric total synthesis of ent‐pyripyropene A was achieved by a convergent synthetic route. We used our originally developed Ti^III‐catalyzed radical cyclization to construct an AB‐ring portion that consisted of a trans‐decalin skeleton with five contiguous stereogenic centers. The coupling between the AB‐ring and the DE‐ring portions, and a subsequent C‐ring cyclization, led to the total synthesis of ent‐pyripyropene A. An evaluation of the insecticidal activity of ent‐pyripyropene A against two aphid species revealed that ent‐pyripyropene A was 35–175 times less active than naturally occurring pyripyropene A. This result indicated that the biological target of pyripyropene A recognizes the absolute configuration of pyripyropene A.     

The structure of neopeptins,inhibitors of fungal cell wall biosynthesis

The structures of neopeptins A (1), B (2), and C (3) were elucidated on the basis of chemical and spectroscopic evidences.     

Studies on the synthesis and biosynthesis of the fungal alkaloid necatorone

An alternative pathway for the biosynthesis of the fungal alkaloid necatorone has been studied using fluorine-labeled 3-(2-carboxyphenylamino)-l-tyrosine. Although no incorporation of this compound could be detected in feeding experiments with young specimens of Lactarius necator, an analogous 3-aminotyrosine derivative could be converted synthetically into the oxopyridoacridine core structure of necatorone. In experiments aimed at the synthesis of aaptamine-type alkaloids, an unprecedented cyclization of a 3-aminotyrosine-methyl propiolate adduct to a methyl isoquinoline-3-carboxylate was observed. A mechanism is proposed, in which C3 of the propiolate delivers C1 of the isoquinoline nucleus.     

Enantioselective Total Synthesis of Terreumols A and C from the Mushroom Tricholoma terreum

The cytotoxic meroterpenoids terreumol A and C from the grey knight mushroom Tricholoma terreum were synthesized for the first time. The key step of the enantioselective total synthesis of terreumol C is a ring‐closing metathesis to form a trisubstituted Z double bond embedded in the 10‐membered ring of the [8.4.0] bicycle. Interestingly, the presence of a free hydroxy group in the metathesis precursor prevents cyclization and favors cross metathesis. (?)‐Terreumol C was converted into (?)‐terreumol A by diastereoselective epoxidation. Starting from 2‐bromo‐3,5‐dimethoxybenzaldehyde, 14 steps with an overall yield of 23 % are needed for the synthesis of (?)‐terreumol A. X‐ray analysis of the benzoquinone analogue of terreumol A provides independent proof of the absolute configuration.     

The biosynthesis of fulvic acid,a fungal metabolite of heptaketide origin

Analyses of the regiochemistry of [¹³C]acetate incorporation into fulvic acid ($\text{2}$) by Penicillium brefeldianum indicates that the metabolite is biosynthesized via a heptaketide intermediate assembled as a single chain of seven C₂, units rather than from two smaller polyketide chains. This information favours a route leading through common C₁₄ intermediates to the three fungal metabolites, citromycetin, fulvic acid and fusarubin.     

Oxidation and biodegradation of polyethylene films containing pro-oxidant additives: Synergistic effects of sunlight exposure, thermal aging and fungal biodegradation

Synergistic effects of sunlight exposure, thermal aging and fungal biodegradation on the oxidation and biodegradation of linear low density poly(ethylene) PE-LLD films containing pro-oxidant were examined. To achieve oxidation and degradation, films were first exposed to the sunlight for 93 days during the summer months followed by their incubation with fungal strains previously isolated from the soil based on the ability to grow on the oxidized PE-LLD as a sole carbon source. Some films were also thermally aged at temperatures ranging between 45°C and 65 °C, either before or after fungal degradation. Films with pro-oxidant additives exhibited a higher level of oxidation as revealed by increase in their carbonyl index (COi). In addition to increase in the COi, films showed a slight increase in crystallinity and melting temperature (T_m), considerably lower onset degradation temperatures, and a concomitant increase in the % weight of the residues. The level of oxidation observed in thermally aged films was directly proportional to the aging temperature. The PE-LLD films with pro-oxidant exposed to sunlight followed by thermal aging showed even higher rate and extent of oxidation when subsequently subjected to fungal biodegradation. The higher oxidation rate also correlated well with the CO₂ production in the fungal biodegradation tests. Similar films oxidized and aged but not exposed to fungal biodegradation showed much less degradation. Microscopic examination showed a profuse growth and colonization of fungal mycelia on the oxidized films by one strain, while another spore-producing strain grew around the film edges. Data presented here suggest that abiotic oxidation of polymer's carbon backbone produced metabolites which supported metabolic activities in fungal cells leading to further biotically-mediated polymer degradation. Thus, a combined impact of abiotic and biotic factors promoted the oxidation/biodegradation of PE-LLD films containing pro-oxidants.     

Guapsidial A and Guadials B and C: Three New Meroterpenoids with Unusual Skeletons from the Leaves of Psidium guajava

A novel sesquiterpene‐based Psidium meroterpenoid, possessing an unusual coupling pattern, and two new monoterpene‐based meroterpenoids with unprecedented skeletons were isolated from the leaves of Psidium guajava. Their structures and absolute configurations were elucidated by spectroscopic, X‐ray diffraction, and computational methods. The plausible biosynthetic pathway of these meroterpenoids as well as their cytotoxicities toward HepG2 and HepG2/ADM cells were also discussed.     

Divergent Total Synthesis of Fornicin A,Fornicin D,and Ganodercin D,Meroterpenoids from Ganoderma Mushrooms

The meroterpenoids fornicin A, fornicin D, and ganodercin D, found in mushrooms of the Ganoderma genus, have been prepared in a concise and divergent synthesis route. The characteristic unsaturated γ-ketoacid moiety was obtained via an optimized step-wise aldol condensation between two readily accessible building blocks. THP-protection of a phenolic hydroxyl group under basic conditions was developed, a protocol that adds to the versatility of this protecting group.