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.


Introduction
Endophytic fungi colonize the internal tissue of plants without causing harm or disease [1]. They can offer protection against predators, pathogens, and abiotic stresses to their host plant [2][3][4][5]. In the past decade, the number of patents has increased to apply endophytic fungi for agricultural, bio-, and phytoremediation purposes, and for the production of active natural products with biomedical applications [6,7].
Endophytic fungi are an important source of active natural products with great chemical diversity which is largely untapped. This biosynthetic capacity and induction for producing secondary metabolites could be related to the activation of genes [8,9]. Many of these compounds possess novel skeletons with antibacterial, antifungal, antiviral, antiinflammatory, antitumor, antimalarial, and other activities, and belong to different classes, such as alkaloids, terpenoids, flavonoids, phenolic compounds, and steroids [10].
Some endophytic fungi can produce the same secondary metabolites of their host plants that are medically important drugs, such as taxol ® , increasing the likelihood of using such endophytes as an alternative and sustainable source for producing these compounds faster than in plants [10].
The production of secondary metabolites by endophytes could be impacted by biotic and abiotic factors, but under lab conditions, due to the selective variation of parameters during cultivation, the culture media and the induction of stress through competition with microorganisms in cocultures represent interesting ways to generate biological activity, chemical diversity, and/or novel uncommon molecules [11,12].
Among the endophytic fungi described in the literature, the genus Aspergillus is the most dominant, representing a rich source of diversity of bioactive natural products [13]. In this review, we cover secondary metabolites from endophytic fungus with uncommon skeletons published from 2015 to 2020. Genera with three or more references comprised the species of Aspergillus, Penicillium, Trichoderma, Chaetomium, Xylaria, Phomopsis, Pestalotiopsis, and Talaromyces. Most of the selected articles showed species characterized through morphological and genetic aspects, mostly using the ITS region and to a lesser extent the 18S region of the ribosomal gene. We use this information to highlight the genetic diversity of endophytes and their ability to produce secondary metabolites with uncommon chemical structures.

Materials and Methods
The search was initially conducted in Scifinder ® using the term "endophytic fungi", and 5942 references were found. Removing duplicates, and refining papers by year (2015-2020), by document type (journal, letter, and review), and by research topic "natural products" resulted in 654 references. Then, we selected 90 articles based on the rareness, uncommonness, unprecedentedly, or novelty on the structure or skeleton of the compounds.
The homologous sequences of the internal transcribed spacer (ITS1) of the ribosomal gene of different species of endophytic fungi were retrieved from the sequence banks of the National Center for Biotechnology Information (NCBI) through the BLAST ® tool (https://blast.ncbi.nlm.nih.gov/Blast.cgi, accessed on 14 July 2021). The search for sequences in the NCBI databases followed the rules recommended by expert mycologists to work with ITS data [14]. The sequences were aligned using the GeneDoc v2.7 program and subsequently analyzed using the neighbor-net phylogenetic-network method with the Kimura 2-parameter model applied in the Splits Tree4 v4.16.2 as previously described (http://nrbsc.org/gfx/genedoc/, accessed on 14 July 2021). Support values were estimated by applying 100 bootstrap replicates. A total of 67 sequences were analyzed, where 32 corresponded to the fungal species described in this review, and 35 were included to represent species of which the genera did not have available sequences in the sequence bank (Table S1). Analysis was also reinforced with a greater representation of taxa.

Results
Since the production of secondary metabolites is dependent on culture settings, around 55% of the articles analyzed here showed the use of rice soaked in distilled water as the main culture medium for fermentation, but it was seeded with a plethora of short-period liquid-culture conditions. Modifications to the solid medium included the addition of natural seawater, saline water, and peptone. Fermentation on a solid rice medium was for long periods (45 days, average) in static conditions in dark or 12/12 h light/dark periods at room temperature. The PDA/PDB medium was the second most used (20%) for fermentation. The rest of the described media were malt extract (MEA), Czapek, and Peptone-Yeast-Glucose (PYG), among others. As opposed to solid fermentations, liquid ones were carried out in rotary shakers at 120 rpm at short periods (15 days, average), controlled pH, and varied conditions of complete dark, light, or 12/12 h dark/light cycles at room temperature.

Indole Alkaloids
Indole alkaloids constitute a group of nitrogen-containing secondary metabolites with interesting chemical structures and diverse biological activities [28]. Figure 2 shows

Indole Alkaloids
Indole alkaloids constitute a group of nitrogen-containing secondary metabolites with interesting chemical structures and diverse biological activities [28]. Figure 2 shows the structures of uncommon indole alkaloids. Giluterrin (24), a prenylated indole alkaloid that presented an unprecedented carbon skeleton, was isolated from endophytic fungus Aspergillus terreus and presented an antiproliferative profile for prostate (PC-3) and kidney (786-0) cancer cell lines [29]. Another rare prenylated indole alkaloid is penioxamide A (25), possessing a piperidine moiety and bearing an antirelative configuration in the bicyclo[2.2.2]diazaoctane ring, and was isolated from Penicillium oxalicum and showed potent brine-shrimp lethality with LD 50 value of 5.6 µM [30].
the structures of uncommon indole alkaloids. Giluterrin (24), a prenylated indole alkaloid that presented an unprecedented carbon skeleton, was isolated from endophytic fungus Aspergillus terreus and presented an antiproliferative profile for prostate (PC-3) and kidney (786-0) cancer cell lines [29]. Another rare prenylated indole alkaloid is penioxamide A (25), possessing a piperidine moiety and bearing an antirelative configuration in the bicyclo[2.2.2]diazaoctane ring, and was isolated from Penicillium oxalicum and showed potent brine-shrimp lethality with LD50 value of 5.6 M [30].

Isoindole Derivatives
Isoindole, a fused benzopyrrole ring system, is the regioisomer of indole heterocycle. Its derivatives have attracted scientific attention for decades, and can be found in natural

Pyrrolidone Derivatives
Pyrrolidine alkaloids are widely distributed in nature and display some interesting biological activities, such antitumor and anti-inflammatory, but microbe-derived ones are rare. Figure 4 shows the structures of uncommon pyrrolidine alkaloids. Collacyclumine A (42) was the first case of a dimeric pyrrolidine alkaloid in nature, and it was isolated from mangrove-derived Colletotrichum salsolae but did not show cytotoxic activity [39]. Paraphaeosphaeride A (43), which possesses a 4-pyranone-γ-lactam-1,4-thiazine moiety, was isolated from endophytic fungus Paraphaeosphaeria neglecta. This compound might provide a new target for synthesis or biosynthetic investigation [40]. Talaramide A (44) is the second example of an alkaloid that possesses a unique oxidized tricyclic system. It was isolated from Talaromyces sp. and displayed promising inhibition of mycobacterial PknG activity [41]. Pericoannosin B (45) was isolated from Periconia sp. and featured the uncommon hexahydro-1H-isochromen-5-isobutylpyrrolidin-2-one skeleton [42].

Pyrrolidone Derivatives
Pyrrolidine alkaloids are widely distributed in nature and display some interesting biological activities, such antitumor and anti-inflammatory, but microbe-derived ones are rare. Figure 4 shows the structures of uncommon pyrrolidine alkaloids. Collacyclumine A (42) was the first case of a dimeric pyrrolidine alkaloid in nature, and it was isolated from mangrove-derived Colletotrichum salsolae but did not show cytotoxic activity [39]. Paraphaeosphaeride A (43), which possesses a 4-pyranone-γ-lactam-1,4-thiazine moiety, was isolated from endophytic fungus Paraphaeosphaeria neglecta. This compound might provide a new target for synthesis or biosynthetic investigation [40]. Talaramide A (44) is the second example of an alkaloid that possesses a unique oxidized tricyclic system. It was isolated from Talaromyces sp. and displayed promising inhibition of mycobacterial PknG activity [41]. Pericoannosin B (45) was isolated from Periconia sp. and featured the uncommon hexahydro-1H-isochromen-5-isobutylpyrrolidin-2-one skeleton [42].

Pyridone and Pyridinyl Derivatives
Pyridone alkaloid asperpyridone A (46) possesses the unusual pyrano[3,2-c]pyridine scaffold and was isolated from Aspergillus sp. This compound demonstrated potential use to develop agents for diabetes treatment due to the pronounced glucose uptake effect on liver HepG2 cells under normal and insulin-resistant conditions [43]. Pair of epimers campyridones A-B (47-48) and C-D (49-50) were isolated from Campylocarpon sp. and featured spiro-furanone or -pyrone moieties, which represent a new family of 4-hydroxy-3-pyridone alkaloids. Only compound 50 showed cytotoxicity against HeLa cells [44].
The 18-hydroxydecaturin B (51) is a novel pyridinyl--pyrone that is rare among natural products, and it was isolated from Penicillium oxalicum and showed brine-shrimp lethality with LD50 value of 2.3 M [30]. Figure 5 shows the structure of uncommon pyridone and pyridinyl derivatives. 3.1.6. Diketopiperazine Derivatives 2,5-Diketopiperazines are cyclopeptides formed by the condensation of two amino acids such as serine, tyrosine, tryptophan, proline, phenylalanine, leucine, isoleucine, histidine, or alanine. They are abundant in nature and are found alone or embedded in larger and complex structures from fungi, bacteria, plants, and mammals [45,46]. Figure  6 shows the structures of uncommon diketopiperazine derivatives.

Pyridone and Pyridinyl Derivatives
Pyridone alkaloid asperpyridone A (46) possesses the unusual pyrano[3,2-c]pyridine scaffold and was isolated from Aspergillus sp. This compound demonstrated potential use to develop agents for diabetes treatment due to the pronounced glucose uptake effect on liver HepG2 cells under normal and insulin-resistant conditions [43]. Pair of epimers campyridones A-B (47-48) and C-D (49-50) were isolated from Campylocarpon sp. and featured spiro-furanone or γ-pyrone moieties, which represent a new family of 4-hydroxy-3-pyridone alkaloids. Only compound 50 showed cytotoxicity against HeLa cells [44].
The 18-hydroxydecaturin B (51) is a novel pyridinyl-α-pyrone that is rare among natural products, and it was isolated from Penicillium oxalicum and showed brine-shrimp lethality with LD 50 value of 2.3 µM [30]. Figure 5 shows the structure of uncommon pyridone and pyridinyl derivatives.

Pyridone and Pyridinyl Derivatives
Pyridone alkaloid asperpyridone A (46) possesses the unusual pyrano[3,2-c]pyridine scaffold and was isolated from Aspergillus sp. This compound demonstrated potential use to develop agents for diabetes treatment due to the pronounced glucose uptake effect on liver HepG2 cells under normal and insulin-resistant conditions [43]. Pair of epimers campyridones A-B (47-48) and C-D (49-50) were isolated from Campylocarpon sp. and featured spiro-furanone or -pyrone moieties, which represent a new family of 4-hydroxy-3-pyridone alkaloids. Only compound 50 showed cytotoxicity against HeLa cells [44].
The 18-hydroxydecaturin B (51) is a novel pyridinyl--pyrone that is rare among natural products, and it was isolated from Penicillium oxalicum and showed brine-shrimp lethality with LD50 value of 2.3 M [30]. Figure 5 shows the structure of uncommon pyridone and pyridinyl derivatives. 3.1.6. Diketopiperazine Derivatives 2,5-Diketopiperazines are cyclopeptides formed by the condensation of two amino acids such as serine, tyrosine, tryptophan, proline, phenylalanine, leucine, isoleucine, histidine, or alanine. They are abundant in nature and are found alone or embedded in larger and complex structures from fungi, bacteria, plants, and mammals [45,46]. Figure  6 shows the structures of uncommon diketopiperazine derivatives.
Epithiodiketopiperazines are cyclic dipeptides containing inter-residual polysulfide bridges between two  carbons. Outrovirin A-C (73-75), gliovirin-like compounds, were identified from Penicillium raciborskii. The sulfide bridge was located between  and  carbons. Compound 75 was the first reported trisulfide gliovirin-like, and it showed antifungal activity against Botrytis cinerea and Verticillium dahliae [54].
Compound trichoderpyrone (82) was isolated from Trichoderma gamsii. It contains a unique cyclopentenone-pyrone hybrid skeleton. This compound displayed weak cytotoxicity against the A-549, HepG2, and HeLa cancer cell lines [57]. Conio-azasterol (83) and S-dehydroazasirosterol (84) were isolated from Coniothyrium cereale, which represented two unusual nitrogen-containing compounds with a sterol portion condensed via two bonds to phenalenone derivatives [58]. Figure 7 shows the structure of other uncommon nitrogen-containing compounds.
Compound trichoderpyrone (82) was isolated from Trichoderma gamsii. It contains a unique cyclopentenone-pyrone hybrid skeleton. This compound displayed weak cytotoxicity against the A-549, HepG2, and HeLa cancer cell lines [57]. Conio-azasterol (83) and S-dehydroazasirosterol (84) were isolated from Coniothyrium cereale, which represented two unusual nitrogen-containing compounds with a sterol portion condensed via two bonds to phenalenone derivatives [58]. Figure 7 shows the structure of other uncommon nitrogen-containing compounds.

Other Types of Alkaloids
The structures of other uncommon alkaloids are shown in Figure 8. The fermentation of endophytic fungus Eupenicillium sp. by epigenetic stimulation led to the enhanced production of new decalin-containing eupenicicols C (85) and D (86), which displayed antimicrobial activity against S. aureus and cytotoxicity against the THP-1 cell line [59]. Penicitroamide (87) was isolated from the culture of Penicillium sp.; its structure contains a bicyclo[3.2.1]octane core with a high degree of carbonylization, and it displayed antibacterial activity against Erwinia carotovora and Sclerotium rolfsii [60]. The liquid culture of Bipolaris sorokiniana resulted in the isolation of isocochlioquinone D (88) and cochlioquinone G (89). Compound 88 featured a rare benzothiazin-3-one moiety, and compound 89 is the first example of cochlioquinones bearing an indole-4,7-dione fragment. Compounds 88 and 89 showed cytotoxic effects against the SF-268, MCF-7, NCI-H460 and HepG-2 cell lines [61]. Phomopsol A (90) was isolated from Phomopsis sp. and presented a highly oxidized polyketide containing the unique 3,4-dihydro-2H-indeno[1,2b]pyridine 1-oxide motif. The compound showed neuroprotective effects against corticosterone-induced injury in PC12 cells [62].

Other Types of Alkaloids
The structures of other uncommon alkaloids are shown in Figure 8. The fermentation of endophytic fungus Eupenicillium sp. by epigenetic stimulation led to the enhanced production of new decalin-containing eupenicicols C (85) and D (86), which displayed antimicrobial activity against S. aureus and cytotoxicity against the THP-1 cell line [59]. Penicitroamide (87) was isolated from the culture of Penicillium sp.; its structure contains a bicyclo[3.2.1]octane core with a high degree of carbonylization, and it displayed antibacterial activity against Erwinia carotovora and Sclerotium rolfsii [60]. The liquid culture of Bipolaris sorokiniana resulted in the isolation of isocochlioquinone D (88) and cochlioquinone G (89). Compound 88 featured a rare benzothiazin-3-one moiety, and compound 89 is the first example of cochlioquinones bearing an indole-4,7-dione fragment. Compounds 88 and 89 showed cytotoxic effects against the SF-268, MCF-7, NCI-H460 and HepG-2 cell lines [61]. Phomopsol A (90) was isolated from Phomopsis sp. and presented a highly oxidized polyketide containing the unique 3,4-dihydro-2H-indeno[1,2-b]pyridine 1-oxide motif. The compound showed neuroprotective effects against corticosterone-induced injury in PC12 cells [62].

Peptides
Four new hybrid peptide-polyketide cyclic depsipeptides, colletopeptides A-D (91-94), were isolated from the culture of endophytic fungus Colletotrichum sp. Their structure featured a rare natural 12-membered cyclic tridepsipeptide containing a 3,5,11-trihydroxy-2-methyl dodecanoic acid unit that represented the first cyclic depsipeptide from the Colletotrichum genus. These compounds exhibited anti-inflammatory activity by inhibiting the production of nitric oxide in RAW264.7 macrophages induced by lipopolysaccharide (LPS); compound 91 also inhibited the production of inflammatory factors IL-6 and TNF- and decreased the phosphorylation of NF-B-associated proteins IB and p65 [63]. Fusarithioamide B (95), a new aminobenzamide derivative, was isolated from Fusarium chlamydosporium and showed selective antifungal activity against C. albicans, and moderate activity against Geotrichum candidum. Moreover, it displayed high antibacterial potential towards E. coli, B. cereus, and S. aureus, a selective and potent effect towards BT-549, MCF-7, SKV-3 and HCT-116, and moderate activity towards the KB and SK-MEL cell lines [64]. Rare depsipeptide chaetomiamide A (96) was isolated from Chaetomium sp. and presented a skeleton with a 13-membered ring system [65]. Unguisin E (97) was obtained from cultures of Mucor irregularis; its structure featured a new -aminobutyric acid-containing cyclic peptide. It did not show biological activity in an antibacterial assay [66]. Figure 9 shows the structures of uncommon peptides.

Peptides
Four new hybrid peptide-polyketide cyclic depsipeptides, colletopeptides A-D (91-94), were isolated from the culture of endophytic fungus Colletotrichum sp. Their structure featured a rare natural 12-membered cyclic tridepsipeptide containing a 3,5,11-trihydroxy-2-methyl dodecanoic acid unit that represented the first cyclic depsipeptide from the Colletotrichum genus. These compounds exhibited anti-inflammatory activity by inhibiting the production of nitric oxide in RAW264.7 macrophages induced by lipopolysaccharide (LPS); compound 91 also inhibited the production of inflammatory factors IL-6 and TNFα, and decreased the phosphorylation of NF-κB-associated proteins IκBα and p65 [63]. Fusarithioamide B (95), a new aminobenzamide derivative, was isolated from Fusarium chlamydosporium and showed selective antifungal activity against C. albicans, and moderate activity against Geotrichum candidum. Moreover, it displayed high antibacterial potential towards E. coli, B. cereus, and S. aureus, a selective and potent effect towards BT-549, MCF-7, SKV-3 and HCT-116, and moderate activity towards the KB and SK-MEL cell lines [64]. Rare depsipeptide chaetomiamide A (96) was isolated from Chaetomium sp. and presented a skeleton with a 13-membered ring system [65]. Unguisin E (97) was obtained from cultures of Mucor irregularis; its structure featured a new γ-aminobutyric acid-containing cyclic peptide. It did not show biological activity in an antibacterial assay [66]. Figure 9 shows the structures of uncommon peptides.

Sesquiterpenoids
The structures of uncommon sesquiterpenoids are shown in Figure 10. A culture of endophytic fungus Zopfiella sp. led to the isolation of undescribed bisabolane sesquiterpenoids zopfiellin B (98) and C (99); the former is a rare trinor-bisabolane sesquiterpenoid, and the latter possesses an unusual aromatic core. These compounds displayed mild cytotoxicity against the A-549, MCF-7, and HeLa cell lines [67]. Aspergoterpenin A (100), isolated from Aspergillus versicolor, is the first example of a ketalbridged-ring part in the degraded natural bisabolane-type sesquiterpene structure. This compound displayed antimicrobial activity against Erwinia carotovora sub sp. [68].

Sesquiterpenoids
The structures of uncommon sesquiterpenoids are shown in Figure 10. A culture of endophytic fungus Zopfiella sp. led to the isolation of undescribed bisabolane sesquiterpenoids zopfiellin B (98) and C (99); the former is a rare trinor-bisabolane sesquiterpenoid, and the latter possesses an unusual aromatic core. These compounds displayed mild cytotoxicity against the A-549, MCF-7, and HeLa cell lines [67]. Aspergoterpenin A (100), isolated from Aspergillus versicolor, is the first example of a ketal-bridged-ring part in the degraded natural bisabolane-type sesquiterpene structure. This compound displayed antimicrobial activity against Erwinia carotovora sub sp. [68]. 021, 7, 570 14 of 31 Figure 10. Structure of sesquiterpenoids.
Two new sesterterpenoids, aspterpenacids A (116) and B (117), featured an unusual pentacarbocyclic 5/3/7/6/5 ring system and were isolated from Aspergillus terreus. Both compounds were tested for antibacterial and cytotoxicity activity, but neither exhibited significant results [78]. Among obtained compounds from the culture of Pseudolagarobasidium acaciicola, acaciicolides A-C (101-103) possess a novel tricyclic ring system and did not show cytotoxic activity [69].

Meroterpenoids
salicyloid derivative containing 2-methylfuran moiety. Compound 143 exhibited in vitro antienterovirus 71 activity that was 5.7 times greater than that of assayed positive-control ribavirin [85]. Asperphenalenones A-E (144-148) were isolated from Aspergillus sp. of the Kadsura longipedunculata plant. Compounds 144-148 represent an unusual linear diterpene derivative linked to a phenalenone derivative via a C-C bond. Compounds 144 and 147 exhibited anti-HIV activity [86]. Another example of a phenalenone derivative was aspergillussanone C (149), obtained from Pinellia ternata endophyte Aspergillus sp. It possesses differentiated acyclic diterpenoid oxidations resulting in an oxa-heterocyclic ring. Compound 149 did not show antimicrobial activity [87].    Figure 13 shows the structures of uncommon polyketides. Phomotide A (150), isolated from endophyte Phomopsis sp., is the first polyketide featuring an unprecedented C12-C6 carbon skeleton. The compound was tested for antibacterial activity but did not show any growth inhibition [88]. A pair of enantiomers, (+)-and (-)-alternarilactone A (151), were isolated from Alternaria sp. and represent the first example of dibenzo--pyrones bearing a diepoxy-cage-like moiety. This pair possessed weak protective activity against corticosterone-induced apoptosis in PC12 cells and did not show significant antibacterial or antifungal effects against some Gram-positive or -negative pathogens [89]. Penicilliumolide A (152) was obtained from the culture of Penicillium chermesinum, and it is a novel tetracyclic polyketide whose structure is uniquely spiro-attached with a -lactone ring. The compound did not show cytotoxicity activity [90].
The culture of Talaromyces wortmanni resulted in the isolation of wortmannin scaffold derivatives, all of which possess an unusual five-membered B ring as in wortmannin A (134); wortmannin B (135) and C (136), additional to ring B, have a lactam and a lactone E-ring, respectively. Compounds 134-136 were evaluated for cytotoxicity but showed no activity [83]. Among compounds isolated from the culture of Guignardia sp., a series of structurally similar meroterpenoids to uncommon tricycloalternarenes (137-142) were characterized to be containing an additional tetrahydrofuran ring. Compound 137 exhibited moderate antibacterial activity against Pseudomonas aeruginosa, while 141 showed prominent inhibition on the growth of C. albicans [84]. Vaccinol J (143) was isolated from the culture of Pestalotiopsis vaccinni, and it represents the first example of salicyloid derivative containing 2-methylfuran moiety. Compound 143 exhibited in vitro antienterovirus 71 activity that was 5.7 times greater than that of assayed positive-control ribavirin [85]. Asperphenalenones A-E (144-148) were isolated from Aspergillus sp. of the Kadsura longipedunculata plant. Compounds 144-148 represent an unusual linear diterpene derivative linked to a phenalenone derivative via a C-C bond. Compounds 144 and 147 exhibited anti-HIV activity [86]. Another example of a phenalenone derivative was aspergillussanone C (149), obtained from Pinellia ternata endophyte Aspergillus sp. It possesses differentiated acyclic diterpenoid oxidations resulting in an oxa-heterocyclic ring. Compound 149 did not show antimicrobial activity [87]. Figure 13 shows the structures of uncommon polyketides. Phomotide A (150), isolated from endophyte Phomopsis sp., is the first polyketide featuring an unprecedented C 12 -C 6 carbon skeleton. The compound was tested for antibacterial activity but did not show any growth inhibition [88]. A pair of enantiomers, (+)-and (-)-alternarilactone A (151), were isolated from Alternaria sp. and represent the first example of dibenzo-α-pyrones bearing a diepoxy-cage-like moiety. This pair possessed weak protective activity against corticosterone-induced apoptosis in PC12 cells and did not show significant antibacterial or antifungal effects against some Gram-positive or -negative pathogens [89]. Penicilliumolide A (152) was obtained from the culture of Penicillium chermesinum, and it is a novel tetracyclic polyketide whose structure is uniquely spiro-attached with a γ-lactone ring. The compound did not show cytotoxicity activity [90].  [62]. Chlorotheolides A (185) and B (186) were isolated from Pestalotiopsis theae, which possesses spiroketals with [4,7]methanochromene and dispirotrione skeletons. Compound 186 showed an antiproliferative effect against HeLa and induced an autophagic process in the cells [105].

Steroids
The structures of uncommon steroids are shown in Figure 14. Phomopsterone A (190), isolated from endophytic fungus Phomopsis sp., is a highly oxygenated ergostane-type steroid possessing a unique bicyclo[3.3.1]nonane motif with an -oriented CH3-19 group. This extremely unusual feature provides new insight into steroid biosynthesis. This compound was tested for in vitro anti-inflammatory activity, but did not show considerable activity [108]. Tricholumin A (191) has an unprecedented carbon skeleton of a highly transformed ergosterol derivative isolated from alga-endophytic Trichoderma asperellum and exhibited inhibition against some pathogenic microbes (V. harveyi, V. splendidus, and Pseudoalteromonas citrea), marine phytoplankton species (Chattonella marina, Heterosigma akashiwo, Karlodinium veneficum, Prorocentrum donghaiense), and antifungal activity against Glomerella cingulata [109]. Secovironolide (192) was isolated from Talaromyces wortmannii and represents the first example of a furanosteroid with a five-membered B-ring in the carbon scaffold. The compound showed weak inhibitory activity in a monoamine oxidase assay [110]. Lanostanoid (193) was isolated from Diaporthe sp. It presented an aromatic B ring with an unusual loss of CH3-19 during the aromatization of the B ring, and hydroxylation at C-1, C-3, C-12, and C-22. This compound is the second reported natural lanostane/cucurbitane derivative with an aromatized B ring. Compound 193 showed pronounced antibacterial efficacy against S. aureus, E. coli, B. subtilis, P. aeruginosa, and S. pyogenes [111]. A pair of enantiomers, (±)-phomone A (153) and (±)-phomone B (154), were isolated from the culture of Phoma sp., and both represent the first examples of 6-α,β-unsaturated ester-2-pyrone dimers; compound 153 also possessed a novel 6/4/5/6 tetracyclic ring system. Both compounds did not show cytotoxicity against three human cancer cells, but the acetylated product of 153 showed moderate cytotoxicity against the HL-60 cell line [91].

Steroids
The structures of uncommon steroids are shown in Figure 14. Phomopsterone A (190), isolated from endophytic fungus Phomopsis sp., is a highly oxygenated ergostane-type steroid possessing a unique bicyclo[3.3.1]nonane motif with an α-oriented CH 3 -19 group.
This extremely unusual feature provides new insight into steroid biosynthesis. This compound was tested for in vitro anti-inflammatory activity, but did not show considerable activity [108]. Tricholumin A (191) has an unprecedented carbon skeleton of a highly transformed ergosterol derivative isolated from alga-endophytic Trichoderma asperellum and exhibited inhibition against some pathogenic microbes (V. harveyi, V. splendidus, and Pseudoalteromonas citrea), marine phytoplankton species (Chattonella marina, Heterosigma akashiwo, Karlodinium veneficum, Prorocentrum donghaiense), and antifungal activity against Glomerella cingulata [109]. Secovironolide (192) was isolated from Talaromyces wortmannii and represents the first example of a furanosteroid with a five-membered B-ring in the carbon scaffold. The compound showed weak inhibitory activity in a monoamine oxidase assay [110]. Lanostanoid (193) was isolated from Diaporthe sp. It presented an aromatic B ring with an unusual loss of CH 3 -19 during the aromatization of the B ring, and hydroxylation at C-1, C-3, C-12, and C-22. This compound is the second reported natural lanostane/cucurbitane derivative with an aromatized B ring. Compound 193 showed pronounced antibacterial efficacy against S. aureus, E. coli, B. subtilis, P. aeruginosa, and S. pyogenes [111].

Analysis of Neighbor Net and Diversity of Endophytic Fungi Producing Secondary Metabolites with Uncommon Structures
The analyzed fungal species in the present review belong to the classes Eurotiomycetes (pink), Sordariomycetes (green), and Dothideomycetes (blue); a minority corresponded to the classes Mucoromycetes, Pinopsida, Agaricomycetes, and Magnoliopsida (Figure 15, yellow). Phylogenetic-network analysis using the internal transcriber spacer (ITS1) of the ribosomal gene indicated that the production of secondary metabolites with uncommon structures is a widely distributed capacity within the seven classes of endophytic fungi analyzed here. Furthermore, endophytic fungal species with wide genetic divergence can generate similar compounds with different uncommon structures. For instance, alkaloids with novel structures were reported in a large number of species in these classes of fungi (Table S1, Figure 15). However, species belonging to the Eurotiomycetes class Aspergillus and Penicillium can biosynthesize a greater variety of these compounds, while the cytochalasin type were mainly isolated from species belonging to the Sordariomycetes class [112,113].

Analysis of Neighbor Net and Diversity of Endophytic Fungi Producing Secondary Metabolites with Uncommon Structures
The analyzed fungal species in the present review belong to classes Eurotiomycetes (pink), Sordariomycetes (green), Dothideomycetes (blue), and a minority corresponds to classes Mucoromycetes and Agaricomycetes (Figure 15, yellow). Phylogenetic-network analysis using the internal transcriber spacer (ITS1) of the ribosomal gene indicated that the production of secondary metabolites with uncommon structures is a widely distributed capacity within the seven classes of endophytic fungi analyzed here. Furthermore, endophytic fungal species with wide genetic divergence can generate similar compounds with different uncommon structures. For instance, alkaloids with novel structures were reported in a large number of species in these classes of fungi (Table S1, Figure 15). However, species belonging to the Eurotiomycetes class Aspergillus and Penicillium can biosynthesize a greater variety of these compounds, while the cytochalasin type were mainly isolated from species belonging to the Sordariomycetes class [112,113].

Discussion
Endophytic fungi producing structurally uncommon compounds listed in the present review were obtained mainly from two hosts: medicinal (37%) and mangrove (24%) plants. These values confirmed the current trend of studying medicinal plantderived-endophytes because it is known that they could have the ability to produce the same metabolites as the host plant [10]. Due to the unique characteristic of the ecosystem, mangrove endophytes have been attractive to the pharma industry, as they produce metabolites that are structurally unique and biologically active [114]. From the 39 different genera reviewed, the majority of references were related to species of Aspergillus (13), Penicillium (11), Trichoderma (6), Phomopsis (5), Chaetomium, Pestalotiopsis, Xylaria (all three with 4) and Talaromyces (3). From the 84 samples of endophytic fungi mentioned in this review, only 57 have been determined to the level of species.
The 202 structures were distributed in six groups: 93 (46%) as alkaloids, 53 (26.2%) as terpenoids, 45 (22.3%) as polyketides, 7 (3.5%) as peptides, and 4 (2%) as steroids. Around 28% were isolated from two main genera, Aspergillus and Penicillium, with 31 and 25 compounds, respectively. For most of these compounds, a biological activity has been determined, including cytotoxic, antitumor, antimicrobial, antiviral, or anti-inflammatory. Table S2 provides information about the endophytes, their host plant, the culture parameters that lead to the production of the uncommon compounds and the biological activity reported.
There is the possibility that these structurally uncommon compounds were produced due to the long culture periods where the fungus starts to consume the first round of produced secondary metabolites to survive starving or a low-resources condition to produce novel and uncommon structures or skeletons via intra-or extramolecular coupling, fusion, or cyclization, such as those described above, in some cytochalasans or polyketides.
Based on the analyzed data, genes mainly involved in the biosynthesis of alkaloids are conserved in a wide range of endophytic fungal species. However, the synthesis of a specific

Discussion
Endophytic fungi producing structurally uncommon compounds listed in the present review were obtained mainly from two hosts: medicinal (37%) and mangrove (24%) plants.
These values confirmed the current trend of studying medicinal plant-derived-endophytes because it is known that they could have the ability to produce the same metabolites as the host plant [10]. Due to the unique characteristic of the ecosystem, mangrove endophytes have been attractive to the pharma industry, as they produce metabolites that are structurally unique and biologically active [114]. From the 39 different genera reviewed, the majority of references were related to species of Aspergillus (13), Penicillium (11), Trichoderma (6), Phomopsis (5), Chaetomium, Pestalotiopsis, Xylaria (all three with 4) and Talaromyces (3). From the 84 samples of endophytic fungi mentioned in this review, only 57 have been determined to the level of species.
The 202 structures were distributed in six groups: 93 (46%) as alkaloids, 53 (26.2%) as terpenoids, 45 (22.3%) as polyketides, 7 (3.5%) as peptides, and 4 (2%) as steroids. Around 28% were isolated from two main genera, Aspergillus and Penicillium, with 31 and 25 compounds, respectively. For most of these compounds, a biological activity has been determined, including cytotoxic, antitumor, antimicrobial, antiviral, or antiinflammatory. Table S2 provides information about the endophytes, their host plant, the culture parameters that lead to the production of the uncommon compounds and the biological activity reported.
There is the possibility that these structurally uncommon compounds were produced due to the long culture periods where the fungus starts to consume the first round of produced secondary metabolites to survive starving or a low-resources condition to produce novel and uncommon structures or skeletons via intra-or extramolecular coupling, fusion, or cyclization, such as those described above, in some cytochalasans or polyketides.
Based on the analyzed data, genes mainly involved in the biosynthesis of alkaloids are conserved in a wide range of endophytic fungal species. However, the synthesis of a specific uncommon structure is probably related to genetic variability and a particular epigenetic aspect of each particular species. Therefore, future research should be focused on the characterization of the biosynthetic gene cluster (BGC), responsible for producing these uncommon structures, to enhance biological synthesis as a tool to produce metabolites of interest. This review demonstrates the importance of endophytic fungi in the production of exclusive metabolites with biotechnological applications. Furthermore, it adequately highlights the value of biological resources for the development of biotrade.