1. Introduction
Endophytic fungi are polyphyletic microorganisms that inhabit plant tissues without inciting disease symptoms, and eventually establish mutualistic associations with their host plants. Until recently these microbial entities have been generally overlooked as a component of ecosystems, the reason why lately they have been regarded as a trove of unexplored biodiversity. Investigations on the biosynthetic potential of endophytes have gained impetus owing to the ongoing discovery of strains capable to synthesize plant compounds, a property which may reflect an adaptative functional role in biocenosis. The intrinsic nature of the interactions among and between endophytes and host plants, and pests, which are mediated by such compounds, is an area open to future discoveries. The elucidation of such connections can not only enhance the understanding of evolution of complex defense mechanisms in plants and their associated organisms, but also help to exploit the latter for a sustained production of a few valuable compounds to be used in biotechnologies [
1].
Secondary metabolites serve in multiple physiologic functions, many of which are common to both plants and microorganisms, and in a way it is intuitive that the same or similar compounds can be produced by ecologically associated entities. Thus, the aim to exploit botanical diversity for the discovery of novel drugs has led to the finding of microbial strains able to synthesize bioactive compounds previously considered as typical plant products. In the last 25 years, the general evidence that all plants are inhabited by endophytic microorganisms, together with an ongoing finding that the latter are also capable of producing plant metabolites, has depicted a research context which is more inclined to consider these compounds as a major factor influencing the establishment and evolution of mutualistic interrelations. Moreover, a refined tool enabling endophytes to regulate the metabolism of host plants in their delicately balanced association [
2].
This paper offers an overview on the extremely varied assortment of organic molecules occurring as secondary metabolites both in plants and endophytic fungi. Our attention is particularly focused on products occurring in single or a restricted range of plant species, with a few exceptions. Therefore, compounds representing metabolic products of a wide array of organisms, such as carbohydrates, aliphatic compounds, aminoacids, peptides, nucleobases, phenolic and benzoic acids, tannins and pseudotannins, sterols, and carotenoids, are not treated in this review.
2. Phytohormones
Plant hormones are undoubtedly among the main secondary metabolites that can influence plant fitness and enhance development when exogenously administered. Many fungal species have been reported to be able to produce compounds such as indole-3-acetic acid (IAA) and gibberellins (GAs), particularly species inhabiting rhizosphere which are presumed to exert a consistent effect on plant development [
3]. Likewise, the observation of such an aptitude by endophytic fungal strains, which in most instances establishes an even closer association with plant tissues, is indicative of a possible functional meaning of mutualistic relationships in certain associations, and provides ground for a more direct impact of an exogenous provision of phytohormones. Unlike what is generally thought for other products synthesized by both plants and endophytes, experimental findings have shown that biosynthetic pathways for these compounds may have evolved independently in plants and fungi [
4,
5].
The interaction of crop plants with endophytic fungi producing gibberellins could be exploited as a strategy to overcome the adverse effects of abiotic stresses, considering the increased plant growth and biomass production that have been documented even in extreme environmental conditions [
6]. On the other hand, it must be considered that hormones are characterized by a dose-related effect, the reason why exceeding supplies may result into abnormal growth. In fact, gibberellins were discovered as products of the plant pathogenic fungus
Fusarium (=
Gibberella)
fujikuroi inducing disease symptoms (elongation of internodes, leaf yellowing,
etc.) in rice plants [
7]. Until recently, a number of fungal plant pathogens have proved to be able to produce these phytohormones as a means for altering physiology of their hosts, but these cases are not a subject for this review. We also do not consider the complex case of the endophytic fungi inhabiting a series of fodder plants, which cause noxious effects to livestock because of their attitude to permeate host tissues with mycotoxins, and are also able to produce phytohormones [
8,
9,
10]. A few excellent reviews on this bordering subject are available in the literature [
11,
12,
13].
Production of IAA by fungi establishing a mutualistic relationship with plants was first evidenced by a few mycorrhizal species recovered from pine and orchid roots [
14,
15,
16]. Afterwards it has been reported by strains of
Colletotrichum sp. from
Artemisia annua [
17],
Talaromyces verruculosus (=
Penicillium verruculosum) from roots of
Potentilla fulgens [
18], and
Penicillium glabrum from pomegranate (
Punica granatum) fruits [
19]. An isolate of the yeast
Williopsis saturnus endophytic in maize roots was found to produce IAA and indole-3-pyruvic acid [
20]. Finally, IAA is also produced by
Piriformospora indica [
21], the quite famous Basidiomycetes species first described as a mycorrhizal agent of shrubs growing in the Rajahstan desert [
22], but later found to be widespread and able to colonize roots of many host plants, disclosing a potential for applications in crop production [
23]. However, all these reports should be carefully verified considering the growth medium used for culturing the producing strains; in fact, it has been observed that the addition of tryptophan incited auxin synthesis by a couple of yeast strains of
Rhodotorula graminis and
R. mucilaginosa recovered from poplar stems [
24].
Other endophytic fungi have been found to produce both IAA and GAs, such as two strains of
Fusarium sp. from
Euphorbia pekinensis [
25]. Moreover, a strain of
Galactomyces geotrichum isolated from the aquatic plant
Trapa japonica produced IAA and biologically active GAs (GA
1, GA
4, and GA
7) [
26], while two strains from cucumber roots (
Phoma glomerata and
Penicillium sp.) were found to produce gibberellic acid (GA
3) along with GA
1, GA
4, GA
7, and IAA in Czapek-Dox broth. This medium is based on sucrose as the only organic compound, which demonstrates an intrinsic ability by these strains to synthesize phytohormones. When experimentally inoculated in cucumber plants under drought stress, the plant biomass and related growth parameters significantly increased, together with a higher assimilation of essential nutrients (K, Ca and Mg), while effects of sodium toxicity were reduced. Moreover, a modulation of stress was also ensured through alteration in jasmonic acid level, down-regulation of abscissic acid, and increased salicylic acid content [
27]. Similar effects resulted in the evaluation of another cucumber endophytic strain (
Paecilomyces formosus) by the same research group [
28].
Production of GAs only has been documented in a higher number of endophytic fungi. A strain of
Fusarium proliferatum from roots of
Physalis alkekengi var.
francheti displayed a plant growth-promoting activity twice stronger than a wild-type of
F. fujikuroi, due to the production of the physiologically active GA
1, GA
3, GA
4, and GA
7, along with GA
9, GA
20 and GA
24 [
29]. Another strain of
F. proliferatum from orchid roots was found to produce GA
4 and GA
7 as the major forms, and smaller amounts of GA
1, GA
3, GA
9, GA
13, GA
14, GA
16, GA
24, GA
25, and GA
36, together with additional side products in the GA-biosynthetic pathway [
30]. A strain of
Scolecobasidium tshawytschae from soybean produced GA
3 together with both active (GA
1, GA
4, and GA
7) and inactive (GA
15 and GA
24) analogs [
31]. Root isolates of
Penicillium citrinum from the dune plant
Ixeris repens [
32], and
Arthrinium phaeospermum from a sedge species (
Carex kobomugi) [
33] both produced GA
1, GA
3, GA
4, and GA
7, along with inactive forms (GA
5, GA
9, GA
12, GA
15, GA
19, GA
20, and GA
24). Strains of
Aspergillus fumigatus,
Cladosporium sphaerospermum and
Talaromyces funiculosus (=
Penicillium funiculosum) from soybean were found to produce bioactive GAs (respectively G
4, G
9, and G
12 the former; GA
3, GA
4, and GA
7 the intermediate; GA
1, GA
4, GA
8, and GA
9 the latter), besides the inactive GA
5, GA
15, GA
19, and GA
24 [
34,
35,
36,
37]. Another strain of
Penicillium sp. from crown daisy (
Chrysanthemum coronarium) was found to promote shoot elongation due to the production of all bioactive GAs (GA
1, GA
3, GA
4, GA
7), along with a few inactive forms (GA
9, GA
12, GA
15, GA
19, GA
20) [
38]. Finally, GA
3 has been reported as a fermentation product of an unidentified endophytic strain from
Curcuma wenyujin [
39].
Cytokinins are a group of
N6-substituted adenine derivatives influencing cell division, vascular development, apical dominance, stress tolerance and leaf senescence. Besides several mycorrhizal fungi reported to contribute to cytokinine levels of roots [
40,
41], production of these phytohormones has been well documented in the above-mentioned
P. indica. In fact, relatively high levels of isopentenyladenine and
cis-zeatins can be found in liquid cultures of this fungus, and accordingly the cytokinine levels are remarkably higher in roots of inoculated plants. Conversely, auxin levels are not influenced despite the ability by
P. indica to produce this phytohormone as well, in connexion with the hypotesized conversion of some IAA into inactive compounds [
42].
Finally, it must be mentioned that production of IAA, GA, abscisic acid and jasmonic acid in several combinations was observed by a pool of unidentified endophytic fungi recovered from five plants used in Indian ethnomedicine,
Camellia caduca,
Osbeckia chinensis,
Osbeckia stellata,
Potentilla fulgens, and
Schima khasiana [
43].
3. Compounds from Essential Oils
Essential oils of plants have a very varied chemical composition, including over 60 different kinds of volatile molecules which can be extracted by distillation and belong to two main groupings, terpenes and aromatic products. Besides contributing to the scent of plants, many of these compounds present interesting bioactive properties, ranging from antibiotic to antitumor effects [
44]. The ability to produce such volatile antibiotics has stimulated to investigate several endophytic strains for their possible use in the so-called mycofumigation of foodstuffs [
45,
46,
47]. Additional prospects for biotechnological applications reside in their use as flavoring agents in the food industry [
48], and even as biofuels [
49]. Not surprisingly, many compounds known from essential oils have been also found as secondary metabolites of endophytic fungi (
Table 1), and novel molecules are more and more characterized as fermentation products of these strains, indicating their effective role in determining the mixture composition.
Table 1.
Components * of essential oils of plants extracted from fungal endophytic strains.
Table 1.
Components * of essential oils of plants extracted from fungal endophytic strains.
Compounds | Producing Species | Host Plants | Reference |
---|
Phenyl propanes |
Asarone | Muscodor tigerii | Cinnamomum camphora | [50] |
Eugenol | Annulohypoxylon stygium | not specified | [51] |
Alternaria sp. | Rosa damascaena | [52] |
Monoterpenes |
Camphor | Nodulisporium sp. | Lagerstroemia loudoni | [47] |
Carene | Meliniomyces variabilis | Pinus sylvestris | [53] |
Nodulisporium sp. | L. loudoni | [47] |
Cineole (eucalyptol) | Nodulisporium sp. | Persea indica | [54] |
L. loudoni | [47] |
Cassia fistula | [55] |
Limonene | unknown Sordariomycetes | Mentha piperita | [56] |
Wickerhamomyces anomalus | Lactuca sativa | [57] |
Nodulisporium sp. | L. loudoni | [47] |
Nigrograna mackinnonii | Guazuma ulmifolia | [58] |
Myrcene | W. anomalus | L. sativa | [57] |
Nodulisporium sp. | L. loudoni | [47] |
Ocimene | W. anomalus | L. sativa | [57] |
Nodulisporium sp. | L. loudoni | [47] |
Phellandrene | Muscodor fengyangensis | Actinidia chinensis, Pseudotaxus chienii | [59] |
Muscodor yucatanensis | Bursera simaruba | [60] |
unidentified strains | “weed grasses” | [61] |
Muscodor sp. | O. granulata | [62] |
Pinane | Nodulisporium sp. | L. loudoni | [47] |
Pinene | unknown Sordariomycetes | M. piperita | [56] |
M. variabilis, Phialocephala fortinii | P. sylvestris | [53] |
W. anomalus | L. sativa | [57] |
N. mackinnonii | G. ulmifolia | [58] |
Sabinanes (thujanes) | Dihydroxysabinane | Phomopsis sp. | Camptotheca acuminata | [63] |
Sabinene (thujene) | Phomopsis sp. | Odontoglossum sp. | [64] |
W. anomalus | L. sativa | [57] |
Nodulisporium sp. | L. loudoni | [47] |
Terpinene, terpineol | Nodulisporium sp. | L. loudoni | [47] |
N. mackinnonii | G. ulmifolia | [58] |
Diterpenes |
Abietadiene | Xylaria sp. | Cupressus lusitanica | [65] |
Totarol | Xylaria sp. | C. lusitanica | [65] |
Sesquiterpenes |
Acoradiene | Phomopsis sp. | Odontoglossum sp. | [64] |
Aristolene | M. yucatanensis | B. simaruba | [60] |
Aromadendrene, isoledene | M. yucatanensis | B. simaruba | [60] |
Phoma sp. | Larrea tridentata | [66] |
Xylaria sp. | C. lusitanica | [65] |
Bisabolene | M. fengyangensis | A. chinensis, P. chienii | [59] |
Xylaria sp. | C. lusitanica | [65] |
Bisabolol | Muscodor kashayum | Aegle marmelos | [67] |
Cadinanes | Cadalenes, calamenenes | Phomopsis cassiae | Cassia spectabilis | [68] |
Bombamalone D, calamenenes, dysodensiol D, indicumolide C | Phomopsis sp. | Pleioblastus amarus | [69] |
Cadinene, amorphene, muurolene | Phoma sp. | L. tridentata | [66] |
Xylaria sp. | C. lusitanica | [65] |
Cubenol | Xylaria sp. | C. lusitanica | [65] |
Carotol | M. tigerii | C. camphora | [50] |
Caryophyllane | Xylaria sp. | C. lusitanica | [65] |
Caryophyllene (humulene) | Muscodor albus | Cinnamomum zeylanicum | [45] |
G. ulmifolia | [70] |
M. fengyangensis | A. chinensis, P. chienii | [59] |
M. yucatanensis | B. simaruba | [60] |
M. variabilis, P. fortinii | P. sylvestris | [53] |
Phoma sp. | L. tridentata | [66] |
Isocaryophyllene | Muscodor sutura | Prestonia trifidi | [71] |
Presilphiperfolanes | Xylaria sp. | Piper aduncum | [72] |
Cedranes | Cedrene, cedrol | Xylaria sp. | C. lusitanica | [65] |
Diepicedrene-1-oxide | M. fengyangensis | A. chinensis, P. chienii | [59] |
Chamigrene | Phoma sp. | L. tridentata | [66] |
M. sutura | P. trifidi | [71] |
Cuparene | unknown Sordariomycetes | M. piperita | [56] |
Drimanes | Albicanol | Perenniporia tephropora | Taxus chinensis var. mairei | [73] |
Hydroxyconfertifolin | Phomopsis sp. | Rhizophora stylosa | [74] |
β-Elemene | Nodulisporium sp. | Cinnamomum loureirii | [46] |
M. yucatanensis | B. simaruba | [60] |
Penicillium baarnense, Penicillium frequentans | Curcuma zedoaria | [75] |
Eremophilanes | Eremophilanolides | Xylaria sp. | Licuala spinosa | [76] |
Isopetasols | unidentified strain | Picea rubens | [77] |
Mairetolide A | Xylaria sp. | C. lusitanica | [78] |
Mairetolide F | Xylaria sp. | L. spinosa | [76] |
Valencene | M. albus | C. zeylanicum | [45] |
unidentified vine plant | [79] |
Oryza granulata | [62] |
Xylaria sp. | C. lusitanica | [78] |
Eudesmanes, eudesmenes (selinenes) | Eutypella sp. | Etlingera littoralis | [80] |
Nodulisporium sp. | C. loureirii | [46] |
Phoma sp. | L. tridentata | [66] |
Xylaria sp. | C. lusitanica | [65] |
Arundinols, arundinones | Microsphaeropsis arundinis | Ulmus macrocarpa | [81] |
Capitulatin B, hydroxycapitulatin B | Nigrospora oryzae | Aquilaria sinensis | [82] |
Farnesene | Xylaria sp. | C. lusitanica | [65] |
Guaianes: guaiene, guaiol (champacol), aciphyllene, bulnesene, gurjunene | M. albus | unidentified vine plant | [79] |
C. zeylanicum | [45] |
Myristica fragrans | [83] |
G. ulmifolia | [70] |
Phoma sp. | L. tridentata | [66] |
Xylaria sp. | C. lusitanica | [65] |
Himachalene | Phoma sp. | L. tridentata | [66] |
Irones | Rhizopus oryzae | Iris germanica | [84] |
Longicyclene | M. variabilis, P. fortinii | P. sylvestris | [53] |
Longifolenes | M. yucatanensis | B. simaruba | [60] |
P. fortinii | P. sylvestris | [53] |
Xylaria sp. | C. lusitanica | [65] |
Longipinene | Phoma sp. | L. tridentata | [66] |
Occidentalol | Xylaria sp. | C. lusitanica | [65] |
Thujopsanol, thujopsene | Xylaria sp. | C. lusitanica | [65] |
M. sutura | P. trifidi | [71] |
Ylangene | Phoma sp. | L. tridentata | [66] |
Zingiberene | Xylaria sp. | C. lusitanica | [65] |
Within the several groups of sesquiterpenes occurring in essential oils, the eremophilanes are particularly widespread among endophytic fungi, and many novel molecules of this series have been first discovered as their fermentation products (
Table 2). It is quite interesting to note that eremophilanes include compounds with phytotoxic effects, such as phaseolinone and phomenone which are also known from an endophytic strain of
Xylaria sp. from the matico tree (
Piper aduncum) [
72].
Table 2.
Eremophilane compounds first characterized from endophytic fungal strains.
Table 2.
Eremophilane compounds first characterized from endophytic fungal strains.
Compounds | Producing Species | Host Plants | Reference |
---|
Botryosphaeridione | Phoma sp. | Melia azedarach | [85] |
Cupressolides A-B | Xylaria sp. | Cupressus lusitanica | [78] |
Dihydroberkleasmin A | Pestalotiopsis photiniae | Podocarpus macrophyllus | [86] |
Eremoxylarins A-B | xylariaceous fungus | not specified | [87] |
MBJ-0011-13 | Apiognomonia sp. | not specified | [88] |
Periconianone B | Periconia sp. | Annona muricata | [89] |
Pestalotiopsin A-B | P. photiniae | P. macrophyllus | [90] |
Phomadecalins | Microdiplodia sp. | Pinus sp. | [91] |
unnamed compounds | Xylaria sp. | Licuala spinosa | [76] |
Xylaria sp. | mangrove plant | [92] |
Xylarenones | Xylaria sp. | Torreya jackii | [93] |
Camarops sp | Alibertia macrophylla | [94,95] |
Data introduced in the above tables are eventually incomplete, due to the ongoing addition of novel findings concerning the direct and indirect influence of endophytic fungi on the composition of essential oils, and ensuing effects on their biological activities. The resulting complex framework and the intriguing applicative perspectives are possibly indicative of the opportunity of a dedicated review for such a hot research topic. A recent one that can be cited here [
96] is limited to sesquiterpenes, and not specifically addressed to fungal endophytes.
Another interesting finding on terpenoids from essential oils concerns a few such compounds, namely aristolene, guaiene and thujopsene, produced by strains of
Daldinia spp. establishing symbiotic relationships with
Xiphydria woodwasps on a number of tree species [
97]. In a strict sense, these fungi cannot be considered endophytic since they are inoculated by the wasps and colonize the galleries excavated by their larvae. However, their belonging to the Xylariaceae, a group including
Xylaria,
Hypoxylon,
Muscodor and
Nodulisporium which are well-known for their endophytic habit, stimulates the appraisal of the ecological role held by these compounds in the identification of the host plant and the establishment of this particular symbiotic relationship, and to consider their possible relevance in other plant-insect interactions.
Finally, the effect of endophytic fungal strains in modifying the pattern of compounds in the essential oils of plants may have intriguing interactions with crop resistance/susceptibility toward pests. As an example, it has been observed that tomato plants harboring an unspecialized endophyte (
Acremonium strictum), which remarkably influences composition of volatile compounds released by the host, are significantly preferred by adult whiteflies (
Trialeurodes vaporariorum) [
98], and for oviposition by the cotton bollworm (
Helicoverpa armigera) [
99]. Therefore, any factors influencing composition of essential oils may be considered in view of enhancing plant protection.
4. Other Terpenoids
Sesquiterpene lactones present some affinity with a few analog components of essential oils, such as guaianes and eudesmanes. Within this category, loliolide (
Figure 1), first characterized from
Lolium perenne [
100], has been reported from a strain of
Annulohypoxylon ilanense from the wood of a plant of
Cinnamomum sp. [
101]. Xanthatin (
Figure 1), first isolated from cockleburs (
Xanthium spp.) and characterized for its antimicrobial, anti-inflammatory, pro-apoptotic and trypanocidal properties [
102,
103], has been recovered from cultures of a strain of
Paecilomyces sp. from
Panax ginseng [
104]. Moreover, dihydrocumambrin A (
Figure 1), known as a secondary metabolite of
Glebionis coronaria (=
Chrysanthemum coronarium), has been recently extracted as a fermentation product of a strain of
Lasiodiplodia (=
Botryodiplodia)
theobromae endophytic in leaves of
Dracaena draco [
105].
Figure 1.
Structures of some terpenoids produced by endophytic fungi.
Figure 1.
Structures of some terpenoids produced by endophytic fungi.
Ginkgolides are diterpenoid lactones known as secondary metabolites of
G. biloba which are considered, particularly ginkgolide B, as prospect drugs based on documented antagonistic effects against the platelet-activating factor involved in a number of inflammatory disorders. Ginkgolides A-C and the related compound bilobalide (
Figure 1) have been found as fermentation products of an endophytic strain of
Fusarium oxysporum recovered from root bark of this living fossil tree [
106].
More terpenoid compounds can be found in oleoresins of certain plants. Agathic acid is a diterpenoid representing the major component of the oleoresin of
Agathis microstachya, an endangered species belonging to the Araucariaceae endemic to Queensland (Australia) [
107], and more recently found in oleoresin of copaiba (
Copaifera spp.) in the Amazon region [
108]. It has been extracted from cultures of a strain of
Botryosphaeria sp. endophytic in leaves of
Maytenus hookeri, together with the related isocupressic acid which is commonly found in the needles of several coniferous species [
109]. These compounds have been reported for an abortive effect on cows [
110], while agathic acid is also known for some extent of trypanocidal activity [
108].
Limonoids are tetranortriterpenes reported from members of Rutaceae and Meliaceae, among which the azadirachtins are best known and exploited for their insecticidal properties [
111]. Azadirachtins A and B (
Figure 2) have been recently extracted from cultures of a strain of
Penicillium (
Eupenicillium)
parvum endophytic in the neem plant (
Azadirachta indica) [
112]. However, the importance of endophytes in the biosynthesis of these plant products may be more substantial, considering that an endophytic
Penicillium strain from the chinaberry tree (
Melia azedarach), which is another source of azadirachtin, has been reported to produce the austins [
113], a series of meroterpenoids which may share the same biosynthetic pathway.
The structurally-related toosendanin (
Figure 2) extracted from fruits and bark of
Melia toosendan, possibly a synonym of
M. azedarach which is used as an anti-helmintic remedy in the Chinese traditional medicine, has been also characterized with reference to its insecticidal effects deriving from antifeedant and growth regulatory properties [
111]. Three unidentified strains endophytic in
M. azedarach have been found to produce this compound [
114], whose possible exploitation as an antitumor drug has been recently envisaged [
115].
Figure 2.
Structures of azadirachtins and toosendanin.
Figure 2.
Structures of azadirachtins and toosendanin.
5. Coumarins
Additional metabolites occasionally found in essential oils, such as bergapten, can be ascribed to the coumarins, a heterogeneous class of natural products whose molecular structure is based on a benzopyran-2-one (
Figure 3). These compounds are widespread in edible plants, therefore commonly assumed through dietary exposure, and considered interesting prospects for a pharmaceutical exploitation due to their antioxidant, anti-inflammatory, anticoagulant, antimicrobial, anticancer, antihypertensive, antihyperglycemic, and neuroprotective properties [
116,
117]. A number of these compounds primarily known from plants have recently started being reported as metabolites of endophytic fungi, such as coumarin found in extracts of strains of
Alternaria spp.,
Penicillium spp. and
Aspergillus flavus from the annual herb
Crotalaria pallida [
118].
Scopoletin (6-methoxy-7-hydroxycoumarin) is reported from several plants, where it is possibly involved in protection against pathogens due to its antimicrobial properties [
119]. This compound has been isolated from a mangrove endophytic strain of
Penicillium sp., together with bergapten and umbelliferone (7-hydroxycoumarin) [
120]. Scopoletin, umbelliferone and 7-
O-prenylumbelliferone, along with additional coumarins known from several plant species such as angelicin, brosiparin, columbianetin, jatamansinol, osthenol and seselin, have been recently extracted from cultures of an endophytic strain of
A. ilanense [
121]. Moreover, marmesin has been reported from a mangrovial endophytic strain of
Fusarium sp. [
122], while bergapten and meranzin have been extracted as fermentation products of a previously-mentioned isolate of
L. theobromae from
D. draco [
105].
Within dihydroisocoumarins, mellein derives its name by a strain of
Aspergillus melleus from which it was first characterized [
123]. It is quite a common fungal metabolite, together with a number of derivatives [
124]. However, several products in this series have been also directly reported from plants [
125,
126]. Examples of compounds produced by both plants and endophytic strains include 7-hydroxymellein extracted from cultures of a strain of
Penicillium sp. associated to
Alibertia macrophylla [
127] and a strain of
Xylaria cubensis derived from leaves of
Litsea akoensis [
128], and mellein itself from a strain of
Nigrospora sp. from the benzoil tree (
Moringa oleifera) [
129]. Another xylariaceous fungus from fruits of the wingleaf soapberry (
Sapindus saponaria) [
130], and a strain of
Annulohypoxylon squamulosum from
Cinnamomum sp. [
131] produced both compounds. Finally, a few mellein derivatives have been found together with isofraxidin (7-hydroxy-6,8-dimethoxycoumarin) as fermentation products of an endophytic strain of
Annulohypoxylon bovei var.
microspora from bark of
Cinnamomum sp. [
132].
Figure 3.
Structures of coumarins produced by endophytic fungi.
Figure 3.
Structures of coumarins produced by endophytic fungi.
6. Flavonoids
Commonly associated to coumarins for their fundamental antioxidant properties, flavonoids also display consistent anti-inflammatory, antibacterial, antiviral, antitumor, and vasodilatatory effects [
133]. These polyphenolic compounds are widespread in plants where they represent a biochemical mechanism in pre- and post-infection resistance against pathogens [
134]. Their structure consists of two benzene rings at either side of a 3-carbon ring, and multiple substitutions in this fundamental frame originate several classes of derivatives, such as flavones, isoflavones, flavonols, flavanones, catechins, and anthocyanins (
Figure 4). Such a variation, and the concomitant occurrence of several compounds belonging to this class is the reason why many reports dealing with their production by endophytic fungal strains consider the total flavonoid content rather than every single product [
135,
136]. When a more detailed analysis is carried out, a complex pattern of metabolites in this series results; this is the case of a strain of
A. ilanense producing kaempferol, quercetin, genkwanin, (+/−)-catechin, (−)-4′-hydroxy-5,7,3′-trimethoxyflavan-3-ol, tectochrysin (5-hydroxy-7-methoxyflavone), dimethylgalangin (3,7-dimethoxy-5-hydroxyflavone), and 5-hydroxy-3,6,7-trimethoxyflavone [
121], and a strain of
Fusarium sp. producing tectorigenin (6-methoxy-5,7,4′-trihydroxyisoflavone) and a few more isoflavones [
122].
Figure 4.
Structures of flavonoids produced by endophytic fungi.
Figure 4.
Structures of flavonoids produced by endophytic fungi.
Kaempferol has been more frequently mentioned as a fermentation product of endophytic fungi, such as a strain of
Mucor fragilis from rhizomes of
Sinopodophyllum hexandrum [
137], a strain of
Fusarium chlamydosporum from stem of
Tylophora indica (Asclepiadaceae) [
138], and a previously-mentioned unidentified strain from
C. wenyujin [
39]. It has been also found together with quercetin in the above-mentioned strain of
A. squamulosum [
131], and in a strain of
A. bovei var.
microspora together with kaempferitrin and luteolin [
139]. A widespread plant metabolite with putative cancer chemopreventive and therapeutic properties [
140], luteolin has been also reported as a secondary metabolite of an endophytic strain of
A. fumigatus from the pigeon pea (
Cajanus cajan), which itself is known as a source of this phytoestrogen [
141].
Cajanol is another isoflavone from roots of
C. cajan displaying antimicrobial properties, and pro-apoptotic activity resulting in assays against human breast cancer cells [
142]. Endophytic strains of
Hypocrea lixii from roots of
C. cajan have also been reported to produce this compound in liquid cultures [
143]. Another endophytic fungus from
C. cajan,
Chaetomium globosum, has been reported to produce apigenin (4′,5,7-trihydroxyflavone) [
144]. Its glycosidic derivative vitexin (apigenin-8-
C-β-
d-glucopyranoside) has been found as a fermentation product of a strain of
Colletotrichum sp. endophytic in
G. biloba [
145]. More glycosidic derivatives, namely apigenin-5-
O-α-
l-rhamnopyranosyl-(1→3)-β-
d-glucopyranoside and euryanoside (apigenin-5-
O-α-
l-rhamnopyranosyl-(1→2)-(6″-
O-acetyl)-β-
d-glucopyranoside), have been detected as transformation products by a strain of
Paraconiothyrium variabile endophytic in the Japanese plum yew (
Cephalotaxus harringtonii) growing on crude extracts of the same plant, by which these compounds had been previously reported [
146].
Rotenone is one of the oldest known bioactive plant metabolites, extracted from roots of tropical Leguminosae such as
Lonchocarpus spp.,
Tephrosia spp.,
Mundulea spp.,
Dalbergia paniculata and
Derris elliptica [
147]. An unidentified
Penicillium strain from the latter plant has been reported to produce this compound or a structural analog [
148].
Silymarin is a bioactive extract of the fruits of milk thistle (
Silybum marianum), containing seven flavolignans with reported antitumor and hepatoprotective properties [
149,
150]. Three of these compounds, silybin A, silybin B, and isosilybin A have been extracted as fermentation products of a strain of
Aspergillus iizukae isolated from the leaves of
S. marianum. Interestingly, flavonolignan synthesis attenuated after repeated subculturing of this strain, but could be resumed when autoclaved leaves of the host plant were added to the growth medium [
151].
The chalcones (benzalacetophenones) are involved in flavonoid biosynthesis in plants through an isomerization process promoted by the enzyme chalcone flavanone isomerase [
152]. These compounds exhibit notable bioactive properties [
153], and significant anti-cancer effects [
154]. An endophytic strain of
Ceriporia lacerata has been found to produce 2′,4′-dihydroxy-6′-methoxy-3′,5′-dimethylchalcone first extracted from its host plant
Cleistocalyx operculatus (=
Eugenia operculata or
Syzygium operculatum) [
155].
7. Xanthones and Quinones
With a number of pharmaceutical applications, xanthones (
Figure 5) are structurally related to flavanoids, and also commonly recovered from both plant and fungal sources [
156]. With reference to endophytic fungi, a mangrovial strain of
Penicillium sp. has been found to produce 1,7-dihydroxyxanthone [
120], primarily known for its antimalarial properties and reported as a secondary metabolite of plants such as
Weddellina squamulosa (Podostemaceae) and several species in the Guttiferae [
157,
158].
Figure 5.
Structures of xanthones produced by endophytic fungi.
Figure 5.
Structures of xanthones produced by endophytic fungi.
Pinselin was initially characterized from a strain of
Penicillium amarum, but later found to be identical to cassiollin reported from
Cassia occidentalis [
159]. This metabolite has been found as a fermentation product of a strain of
Phomopsis sp. endophytic in rhizome of
Paris polyphylla var.
yunnanensis [
160], together with additional xanthones including 6-
O-methyl-2-deprenylrheediaxanthone B, previously extracted from bark of
Garcinia vieillardii [
161].
Fairly widespread as plant metabolites and widely used as dyes in the textile industry, anthraquinones are believed to contribute to the defense against pests and disease agents, based on their antifungal, antibacterial and insecticidal properties [
162]. Several compounds in this class are also known from endophytic fungi (
Figure 6). Emodin, physcion and citreorosein, found in several plant species such as rhubarb (
Rheum spp.) [
163],
Cassia spp. [
164], and
Polygonum cuspidatum [
165,
166], are also produced by heterologous endophytic strains of
Penicillium spp. (e.g.,
P. herquei,
P. janthinellum) [
167,
168,
169]. Emodin has been also reported from a strain of
Talaromyces sp. from the mangrove
Kandelia candel [
170], while physcion was also extracted from cultures of an unidentified strain from the mangrove
Avicennia marina [
171], and of strains of
A. fumigatus from
Cynodon dactylon [
172], and
Aspergillus terreus from
Opuntia ficus-indica [
173]. Quinizarin known from roots of the madder plant (
Rubia tinctorum) [
174], has been reported as a secondary metabolite of a previously-mentioned unidentified strain from
C. wenyujin [
39]. Endocrocin, reported from roots of
Rumex nepalensis [
175], has recently been found to be produced by an endophytic strain of
Pestalotia (
Pestalotiopsis)
acaciae [
176]. Questinol and questin are known from
Polygonum spp. [
177] and again from
Cassia spp. [
164]. The former compound has been detected in the culture extracts of strains of
Eurotium rubrum from the mangrove
Hibiscus tiliaceus [
178] and
P. glabrum from pomegranate fruits [
19], while the latter has been reported as a metabolite of an unidentified strain belonging to the Dothideomycetes from the Thai medicinal plant
Leea rubra. This strain also produces the biosynthetically related chromone derivative eugenitin, previously extracted as a secondary metabolite of clove (
Syzygium aromaticum), and its analog 6-hydroxymethyleugenitin [
179]. A strain of
Phoma sorghina endophytic in the medicinal plant
Tithonia diversifolia [
180] has been found to produce pachybasin and phomarin, known as secondary metabolites of species in the Scrophulariaceae, such as
Digitalis spp. [
181] and
Isoplexis isabelliana [
182], and 1,7-dihydroxy-3-methylanthraquinone, first extracted from leaves of
Digitalis viridiflora [
183]. Once again, the three latter compounds have been more recently extracted from cultures of a strain of
Coniothyrium sp. endophytic in
Salsola oppostifolia [
184]. Finally, pachybasin has been also reported from an unidentified strain from the yellow moonshed (
Arcangelisia flava) [
185].
Hypericin, a secondary metabolite of St. John’s wort (
Hypericum perforatum) and other congeneric species, has been extensively studied for its antidepressant, antiviral and antitumor properties. In combination with light the compound effectively induces apoptosis and/or necrosis of cancer cells, the reason why it has been exploited for the photodynamic therapy of cancer [
186]. Hypericin has been found as a metabolic product of an endophytic strain of
Thielavia subthermophila isolated from
H. perforatum in India, together with the above-mentioned emodin considered as a possible precursor [
187,
188].
Figure 6.
Structures of anthraquinones produced by endophytic fungi.
Figure 6.
Structures of anthraquinones produced by endophytic fungi.
Sterequinone C, primarily extracted from bark of the Ayurvedic medicinal plant
Stereospermum personatum [
189], has been more recently found as a metabolite of an unidentified strain of
Penicillium sp. from leaf of the mangrove
Avicennia [
120].
The tanshinones are diterpenoid quinones from roots and rhizomes of danshen (
Salvia miltiorrhiza) and other
Salvia spp. exerting antioxidant, anti-inflammatory and antitumor effects [
190]. These compounds have been also extracted from endophytic strains of the same plant, particularly TR21 of
Emericella foeniculicola [
191], and D16 of
Trichoderma atroviride [
192]. Polysaccharides from the mycelium of the latter strain were found to elicit root growth and tanshinone production [
193]. Moreover, a previously-mentioned strain of
Paecilomyces sp. from
P. ginseng has been found to produce isotanshinone [
104].
Shikonin is a pigment of the Chinese therapeutic herb zicao (
Lithospermum erythrorhizon) with a naphthoquinone structure, displaying interesting pharmacological activities based on its anti-inflammatory, antigonadotropic, anti-HIV and antitumor properties [
194]. This compound has been recently recovered from extracts of an endophytic strain from roots of the old lady cactus (
Mammillaria hahniana) [
195]. Lapachol is another naphthoquinone derivative originally extracted from the bark of the lapacho tree (
Handroanthus impetiginosus, syn.
Tabebuia avellanedae), but later found in plants belonging to different botanical families. It is an active principle in lapacho used in ethnomedicine by indigenous populations of South America, and reported for a wide range of pharmacological activities [
196]. Endophytic strains of
Alternaria sp.,
A. alternata,
A. niger and
Penicillium sp. from the silver trumpet tree (
Tabebuia argentea) have been found to produce this compound [
197,
198].
8. Lignans
Quite widespread as plant metabolites [
199], the podophyllotoxins and related cyclolignans have been also characterized for their insecticidal effects [
200,
201]. However, the higher reputation for podophyllotoxin (
Figure 7) and a few synthetic derivatives relies on their pharamacological applications, particularly as antitumor drugs [
199,
202]. These products have also been reported from an increasing number of endophytic fungi, such as
Alternaria sp. from
S. hexandrum [
203] and
Juniperus communis (=
J. vulgaris) [
204],
Penicillium spp. from
S. hexandrum,
Diphylleia sinensis and
Dysosma veitchii [
203],
Phialocephala fortinii from
Podophyllum peltatum [
205],
Trametes hirsuta,
Fusarium solani and
M. fragilis from
S. hexandrum [
137,
206,
207], and
F. oxysporum from
Juniperus recurva [
208]. Moreover, deoxypodophyllotoxin has been reported from a strain of
A. fumigatus endophytic in
J. communis [
209].
Phillyrin (
Figure 7) is another lignan displaying antioxidant, anti-inflammatory and antipyretic activities, known from
Phyllirea and a number of medicinal plants such as
Forsythia suspensa, a shrub used in Chinese traditional medicine [
210]. An endophytic strain of
Colletotrichum gloeosporioides from a fruit of this plant was found to produce phillyrin in liquid cultures [
211]. Other lignans known from several plant species, such as sesamin, syringaresinol and ketopinoresinol (
Figure 7), have been recently found as secondary metabolites of an endophytic strain of
A. ilanense [
121].
Figure 7.
Structures of lignans produced by endophytic fungi.
Figure 7.
Structures of lignans produced by endophytic fungi.
9. Taxol and Taxanes
Taxol, also known as paclitaxel, is probably the most valuable compound in this review, at least considering its notoriety as the first billion-dollar anticancer drug. Temporarily set aside in chapter 4 concerning terpenoids, this diterpene compound presenting an unusual oxytane ring and a tricyclic core (
Figure 8) was originally characterized from bark of the Pacific yew (
Taxus brevifolia) [
212], but later found in all other yew species [
213]. Problems for an adequate pharmaceutical supplying were soon evident, and the need to find alternative sources lead to the discovery of the first endophytic fungal strain able to synthesize this compound, isolated from the inner bark of
T. brevifolia and ascribed to the novel species
Taxomyces andreanae [
214]. This discovery was followed by an ongoing series of similar findings from yews, but also from other plants species, attaining to a provisional number of almost 100 reports of endophytic strains belonging to 72 fungal species from 32 different host plants, as listed in
Table 3. However, these figures are underestimated, considering that we have not been able to check a number of reports from China and other Asian countries. Whether or not one or more of such strains can be effectively employed by the pharmaceutical industry is still under debate [
1,
215], and treating this theme goes beyond the scopes of this review. Nevertheless, there is no doubt that the issue of taxol production by endophytic fungi has stimulated a wide research activity concerning drugs and other reputed bioactive compounds extracted from plants, introducing new insights in the appreciation of the microbial component of biodiversity and the opportunity of its exploitation for human wellness.
Figure 8.
Structures of taxol and baccatins.
Figure 8.
Structures of taxol and baccatins.
Table 3.
Endophytic fungi reported for production of taxol.
Table 3.
Endophytic fungi reported for production of taxol.
Species | Host plants | Reference |
---|
Acremonium sp. | Taxus globosa | [216] |
Alternaria sp. | Taxus cuspidata Ginkgo biloba Corylus avellana | [217] [218] [219] |
Aspergillus candidus | Taxus × media | [220] |
Aspergillus fumigatus | Podocarpus sp. | [221] |
Aspergillus niger var. taxi | T. cuspidata | [222] |
Aspergillus sp. | Taxus chinensis | [223] |
Bartalinia robillardoides | Aegle marmelos | [224] |
Botryosphaeria sp. | T. globosa | [216] |
Botrytis sp. | T. chinensis var. mairei T. cuspidata | [225] [226] |
Ceratobasidium sp. | T. chinensis | [223] |
Chaetomella raphigera | Terminalia arjuna | [227] |
Cladosporium cladosporioides | T. media | [228] |
Cladosporium oxysporum | Moringa oleifera | [229] |
Cladosporium tenuissimum | T. chinensis | [223] |
Colletotrichum gloeosporioides | Justicia gendarussa Plumeria acutifolia T. media Tectona grandis M. oleifera | [230] [231] [232] [233] [234] |
Colletotrichum sp. | Maguireothamnus speciosus | [235] |
Coniothyrium diplodiella | T. chinensis | [223] |
Didymostilbe sp. | T. chinensis var. mairei | [236] |
Ectostroma sp. | T. chinensis var. mairei | [225] |
Epacris sp. | T. chinensis | [223] |
Fusarium arthrosporioides | T. cuspidata | [237] |
Fusarium lateritium | Taxus baccata | [217] |
Fusarium mairei | T. chinensis var. mairei | [238] |
Fusarium oxysporum | Rhizophora annamalayana | [239] |
Fusarium proliferatum | T. media | [232] |
Fusarium redolens | Taxus wallichiana | [240] |
Fusarium solani | Taxus celebica T. chinensis | [241] [242] |
Fusarium sp. | T. wallichiana T. globosa | [243] [216] |
Gliocladium sp. | T. baccata | [244] |
Guignardia mangiferae | T. media | [232] |
Gyromitra sp. | T. globosa | [216] |
Lasiodiplodia theobromae | Morinda citrifolia T. baccata | [245] [246] |
Metarhizium anisopliae | T. chinensis | [223] |
Monochaetia sp. | T. baccata | [217] |
Mucor rouxianus | T. chinensis | [247] |
Mucor sp. | T. chinensis var. mairei | [248] |
Nigrospora sp. | T. globosa | [216] |
Nodulisporium sylviforme | T. cuspidata | [249] |
Ozonium sp. | T. chinensis var. mairei | [250] |
Paraconiothyrium brasiliense | T. chinensis | [223] |
Paraconiothyrium sp. | T. media | [251] |
Penicillium aurantiogriseum | C. avellana | [252] |
Penicillium raistrickii | Taxus brevifolia | [253] |
Penicillium sp. | Taxus yunnanensis (=T. wallichiana) T. globosa T. chinensis | [254] [216] [255] |
Periconia sp. | Torreya grandifolia | [256] |
Pestalotia bicilia | T. baccata | [217] |
Pestalotia pauciseta | Cardiospermum helicacabum Tabebuia pentaphylla | [257] [258] |
Pestalotiopsis guepinii | Wollemia nobilis | [259] |
Pestalotiopsis microspora | T. cuspidata, T. wallichiana Taxodium distichum | [260] [261] |
Pestalotiopsis neglecta | T. cuspidata | [262] |
Pestalotiopsis sp. | W. nobilis Catharanthus roseus | [259] [263] |
Pestalotiopsis terminaliae | T. arjuna | [264] |
Pestalotiopsis versicolor | T. cuspidata | [262] |
Pezicula sp. | T. chinensis | [223] |
Phoma betae | G. biloba | [265] |
Phomopsis sp. | G. biloba, Larix leptolepis, T. cuspidata T. chinensis | [266] [223] |
Phyllosticta melochiae | Melochia corchorifolia | [267] |
Phyllosticta sp. | Ocimum basilicum | [268] |
Phyllosticta spinarum | Cupressus sp. | [269] |
Pithomyces sp. | Taxus sumatrana | [216] |
Rhizopus sp. | T. media | [270] |
Seimatoantlerium nepalense | T. wallichiana | [271] |
Sordaria sp. | T. chinensis | [223] |
Sporormia minima | T. wallichiana | [272] |
Stegolerium kukenani | Stegolepisguianensis | [273] |
Stemphylium sedicola | T. baccata | [274] |
Taxomyces andreanae | T. brevifolia | [214] |
Taxomyces sp. | Taxus sp. | [275] |
Trichoderma sp. | T. chinensis | [223] |
Trichothecium sp. | T. wallichiana | [272] |
Tubercularia sp. | T. mairei | [276] |
Xylaria sp. | M. speciosus T. chinensis | [235] [223] |
Another opportunity to obtain taxol is represented by a semisynthetic method starting from the structurally related taxane compounds baccatin III and 10-deacetylbaccatin III (
Figure 8), which are more easily accessible for their occurrence in yew needles [
277]. Besides a number of papers where these precursors are reported together with taxol (
Table 3), specific references consider their production by endophytic strains from
Taxus wallichiana, respectively
Diaporthe phaseolorum [
278] and
Trichoderma sp. [
279]. A more general biosynthetic ability concerning taxanes has been evidenced in a number of endophytic strains isolated from
Taxus baccata which were not identified at the species level, ascribed to the genera
Alternaria,
Aspergillus,
Beauveria,
Epicoccum,
Fusarium,
Gelasinospora,
Geotrichum,
Phoma and
Phomopsis [
280]. Evidence as taxane producers has been also reported for strains of
Cladosporium langeronii and
Phomopsis sp. from
Wollemia nobilis [
281].
10. Quinoline Alkaloids
Camptothecin (CPT) is probably the best known representative of this group, whose importance and history as a plant-derived antitumor drug to be widely found as a secondary metabolite of endophytic fungi seems to retrace what just described for taxol. In fact, in the last decade, this compound, and its analogues 9-methoxycamptothecin (MCPT) and 10-hydroxycamptothecin (HCPT) (
Figure 9), have been reported from a number of endophytes from
Camptotheca acuminata and a few more trees in the unrelated botanical family of the Icacinaceae (
Table 4).
The attempt to obtain CPT by fermentation has evidenced a fundamental problem which may affect the possible use of endophytic fungal strains for production of bioactive plant metabolites on a large scale. In fact, the loss of the biosynthetic ability has been documented in a number of strains of
F. solani [
282] and
Aspergillus sp. [
283]. This breakdown seems not to be related to the antibiotic properties of the compound, considering that resistance to CPT has been documented for a number of endophytic strains from
C. acuminata [
284], and it is intrinsic and not related to the biosynthetic aptitude [
285]. On the other hand, production of camptothecins can be stimulated by exploiting the eliciting effects of a few organic and inorganic compounds, particularly salicylic acid which induced a higher HCPT yield by a strain of
Xylaria sp. [
286], and methyl jasmonate which increased CPT production by a strain of
T. atroviride [
283]. Furthermore, CPT production by a strain of
F. solani from
C. acuminata was notably increased by supplying an ethanolic extract of leaves of
C. roseus containing strictosidine as a precursor in CPT biosynthesis, and even by ethanol itself [
287].
Figure 9.
Structures of quinoline alkaloids produced by endophytic fungi.
Figure 9.
Structures of quinoline alkaloids produced by endophytic fungi.
Table 4.
Endophytic fungi producing camptothecin (CPT) and its analogues 9-methoxycamptothecin (MCPT) and 10-hydroxycamptothecin (HCPT).
Table 4.
Endophytic fungi producing camptothecin (CPT) and its analogues 9-methoxycamptothecin (MCPT) and 10-hydroxycamptothecin (HCPT).
Compounds | Fungal Species | Host Plants | References |
---|
CPT | Entrophospora infrequens | Nothapodytes foetida | [288,289] |
CPT | Neurospora sp. | N. foetida | [290] |
CPT | Nodulisporium sp. | N. foetida | [291] |
CPT, HCPT, MCPT | Fusarium solani | Camptotheca acuminata | [292] |
CPT, HCPT, MCPT | F. solani | Apodytes dimidiata | [293] |
CPT | Botryosphaeria parva | Nothapodytes nimmoniana | [294] |
CPT | Diaporthe conorum | N. nimmoniana | [294] |
CPT | Fusarium oxysporum | N. nimmoniana | [294] |
CPT | Fusarium sacchari | N. nimmoniana | [294] |
CPT | F. solani | N. nimmoniana | [294] |
CPT | Fusarium sp. | N. nimmoniana | [294] |
CPT | Fusarium subglutinans | N. nimmoniana | [294] |
CPT | Fusarium verticillioides | N. nimmoniana | [294] |
CPT | Galactomyces sp. | N. nimmoniana | [294] |
CPT | Irpex lacteus | N. nimmoniana | [294] |
CPT | Phomopsis sp. | N. nimmoniana | [294] |
CPT | Unidentified strains | N. nimmoniana | [294] |
HCPT | Xylaria sp. | C. acuminata | [286] |
HCPT | Valsa mali | C. acuminata | [295] |
CPT | Aspergillus spp. | C. acuminata | [283] |
CPT | Trichoderma atroviride | C. acuminata | [283] |
CPT, HCPT, MCPT | Alternaria alternata | Miquelia dentata | [296] |
CPT, HCPT, MCPT | Fomitopsis sp. | M. dentata | [296] |
CPT, HCPT, MCPT | Phomopsis sp. | M. dentata | [296] |
CPT | F. oxysporum | N. foetida | [297] |
A paramount pharmaceutical relevance also pertains to the cinchona-alkaloids (
Figure 9) extracted from bark of quina trees (
Cinchona sp.), representing fundamental drugs for malaria prophylaxis [
298]. After reporting the first endophytic isolate (
Diaporthe sp.) from
Cinchona ledgeriana collected in Java (Indonesia) producing quinine, quinidine, cinchonidine, and cinchonine [
299], additional 20 strains of
Phomopsis sp.,
Schizophyllum sp.,
Penicillium sp.,
Fomitopsis sp., and
Arthrinium sp. from the same source have been found to produce variable amounts of the these compounds [
300,
301].
Two more compounds in this class are to be considered in this review. Berberine (
Figure 9) is a cardioprotective, antidiabetic, antibiotic and antitumor product known from several unrelated medicinal plants [
302], which has been recently found as a secondary metabolite of a strain of
F. solani from roots of the medicinal liana
Coscinium fenestratum [
303]. Sanguinarine (
Figure 9), a benzylisoquinoline or benzophenanthridine alkaloid known from several plants belonging to the Papaveraceae, whose antimicrobial effects have been particularly exploited for toothpastes and mouthwashes [
304], has been found to be produced by a strain of
F. proliferatum from leaves of
Macleaya cordata [
305].
11. Other Alkaloids
The vinca-alkaloids (
Figure 10) are a series of over a hundred bioactive products extracted from periwinkle (
Catharanthus roseus) and the related
Vinca species, mostly exploited in cancer chemotherapy based on their action on tubulin and microtubule organization [
306]. Endophytic strains of
Alternaria sp. [
307] and
F. oxysporum [
308] from
C. roseus respectively produced vinblastine and vincristine, while more recently another strain of the latter species has been found to produce both compounds [
309], and to synthesize vincristine from vinblastine when the latter compound was added to the growth medium [
310]. Finally, an unidentified strain from
Vinca minor has been reported for production of vincamine [
311].
Figure 10.
Structures of vinca-alkaloids.
Figure 10.
Structures of vinca-alkaloids.
Caffeine, the methylxanthine alkaloid (
Figure 11) from
Coffea spp. well known as a psychoactive drug, has been evidenced in extracts of unidentified endophytic fungi recovered from the Indian ethnomedicinal plants
Osbeckia chinensis,
O. stellata and
Potentilla fulgens [
43].
Piperine (
Figure 11) is an alkaloid of
Piper longum and
Piper nigrum known for its antimycobacterial, antihyperlipidemic, anti-inflammatory, immunoregulatory and antitumor properties [
312,
313,
314]. This valuable compound has been extracted from liquid cultures of endophytic strains of
Periconia sp. from
P. longum [
315], and
Mycosphaerella sp. [
316] and
C. gloeosporioides [
317] from
P. nigrum.
Figure 11.
Structures of miscellaneous alkaloids produced by endophytic fungi.
Figure 11.
Structures of miscellaneous alkaloids produced by endophytic fungi.
Aconitine is a diterpenoid alkaloid (
Figure 11) extracted as a secondary metabolite of
Aconitum species, displaying analgesic, anti-inflammatory and antitumor activity. It has been also found as a fermentation product of an endophytic strain of
Cladosporium cladosporioides recovered from roots of
Aconitum leucostomum [
318].
Rohitukine is a chromane alkaloid (
Figure 11) with anti-inflammatory, immunomodulatory and antitumor properties first characterized as a secondary metabolite of a few tropical plants,
Amoora rohituka (=
Aphanamixis polystachya) and
Dysoxylum binectariferum (Meliaceae),
Schumanniophyton (
Tetrastigma)
magnificum and
Schumanniophyton (
Assidora)
problematicum (Rubiaceae) [
319]. This compound has been recently reported as a fermentation product of endophytic strains of
F. proliferatum,
F. oxysporum and
F. solani from
D. binectariferum [
320,
321], and
F. fujikuroi from
A. rohituka [
321].
Peimisine and peiminine are steroidal alkaloids (
Figure 11) known to occur in bulbs of
Fritillaria spp. used as in Chinese ethnomedicine for their expectorant effects. A strain of an undetermined
Fusarium species endophytic in bulbs of
Fritillaria unibracteata var.
wabensis has been found to produce these compounds [
322].
Swainsonine is a trihydroxy inolidizine alkaloid (
Figure 11) acting as a glycosidase inhibitor which occurs in a number of plant species, such as
Swainsona canescens,
Ipomoea spp.,
Turbina cordata,
Sida carpinifolia, and locoweeds (
Astragalus and
Oxytropis spp.) [
323]. The latter are forage plants whose consumption by livestock is associated to the syndrome of locoism. It has been observed that a fungal endophytic species occurring in locoweeds, the vertically-transmitted
Undifilum oxytropis [
324,
325], previously described as
Embellisia sp. [
326,
327], is capable to produce swainsonine
in vitro. Novel species of
Undifilum,
U. fulvum and
U. cinereum, have been later reported for production of this compound, respectively in
Astragalus lentiginosus and
A. mollissimus [
328], along with strains of an unidentified
Undifilum sp. recovered from
S. canescens in Australia [
329]. Moreover, the compound was found to be produced by endophytic isolates of
Fusarium tricinctum recovered from
Oxytropis kansuensis and
O. deflexa in China [
330]. The inhibitory properties on α-mannosidase have been exploited for the development of swainsonine as an anticancer drug, which has been impaired by reason of undesirable side effects [
331,
332]. Finally, it is interesting to note that plants of
A. lentiginosus and
Oxytropis sericea germinated from the embryo, which is not colonized by endophytes, were found to be fungus free and did not contain swainsonine [
333]. This evidence strongly supports the hypothesis that actually its occurrence in plants may entirely derive from biosynthesis by endophytic fungal strains.
13. Saponines
Saponines are glycosides where the sugar moiety is bound through a glycosidic linkage to an aglycone (sapogenin) which can be a triterpenoid or a steroid compound. Generally considered as antinutritional factors, these products have been recently re-evaluated for their nutraceutical properties deriving from consistent anticholesterolemic effects and inhibition of sugar and ethanol absorption. Additional antibiotic, antitumor and immunomodulatory properties have also been reported [
342]. Saponins are known to occur in many taxonomically unrelated plants, but there is an increasing evidence that their production is also widespread among endophytic fungi. Again such a biosynthetic ability may represent an aspect of the mutualistic relationships established in view of plant defense, considering that some plant pathogens are reported to have developed saponin-detoxifying enzymes as a virulence factor [
343]. To this regard, production of saponins has been recently documented for strains of
Aspergillus sp. from
Salvadora oleoides [
344] and
Justicia beddomei [
345],
Aspergillus sp.,
Bulgaria sp. and
Sirococcus conigenus from
Potentilla fulgens [
346],
A. niger and
F. oxysporum from
Crotalaria pallida [
347],
A. alternata,
A. niger and
Penicillium sp. from
Loranthus sp. [
348],
A. alternata,
A. flavus,
A. niger,
Colletotrichum gleosporioides and
Trichoderma sp. from
Tabebuia argentea [
197],
Cochliobolus lunatus (anamorph
Curvularia lunata) from
Boswellia ovalifoliolata, and
Monochaetia karstenii (=
Pestalotiopsis maculans) and
Phyllosticta sp. from
Shorea thumbuggaia [
349],
Aspergillus neoniveus (=
Fennellia nivea) from
Typhonium divaricatum [
350],
A. alternata,
A. flavus,
A. niger,
Cladosporium sp.,
Penicillium sp.,
Phomopsis sp. and
Trichoderma sp. from
Aegle marmelos [
351],
A. niger,
A. terreus,
Aspergillus sp.,
Aspergillus tubingensis,
Coprinopsis cinerea,
C. lunata and
Fusarium sp. from
Eugenia jambolana [
352],
Aspergillus awamori and again
C. gleosporioides from
Rauwolfia serpentina [
353].
More particularly, the ginsenosides (
Figure 12), previously characterized from plants in the genus
Panax, have been extracted from cultures of root endophytic strains of
Penicillium sp.,
Dictyochaeta sp. and
Camarosporium sp. from
Aralia elata [
354], and of
Fusarium sp.,
Aspergillus sp. and
Verticillium sp. from
P. ginseng [
355]. However, the ability by endophytic strains of
F. oxysporum,
Fusarium sp. and
Nodulisporium sp. from
Panax notoginseng to transform ginsenosides to yield additional analog compounds [
356] demonstrates that endophytic fungi directly contribute to the particular pattern of these compounds occurring in plant tissues.
Figure 12.
Structures of saponines produced by endophytic fungi.
Figure 12.
Structures of saponines produced by endophytic fungi.
A similar effect has been documented for an endophytic strain of
Fusarium sp. from
Dioscorea nipponica which was able to increase diosgenin content in its liquid cultures added with a rhizome extract of the host plant [
357]. Diosgenin is a steroidal sapogenin (
Figure 12) known as a secondary metabolite of a number of plant species, particularly the yams (
Dioscorea spp.). It is an important precursor for production of semi-synthetic steroids, such as corticosteroids, progesterone and other steroidal drugs, and displays a number of important pharmacological effects [
358]. This valuable compound is also produced by endophytic strains of
Cephalosporium sp. and
Paecilomyces sp. from
P. polyphylla var.
yunnanensis [
359,
360].
In vitro cell culturing represents an alternative means for production of this drug, considering that natural populations of one of the most important sources, the yellow ginger (
Dioscorea zingiberensis), are decreasing due to overexploitation, and agricultural production is problematic by reason of the 3–4 years required for obtaining mature rhizomes. Oligosaccharides and polysaccharides extracted from an endophytic strain (Dzf17) of
F. oxysporium have been found to elicit growth and diosgenin production in cell cultures of
D. zingiberensis [
361,
362]. The same effect was also observed when both cell cultures and seedlings were treated with beauvericin produced by an endophytic strain of
Fusarium redolens from the same plant species [
363].
Gymnemagenin, a triterpenoid sapogenin (
Figure 12) extracted from the renowned anti-diabetic herb
Gymnema sylvestre, has been found as a fermentation product of an endophytic strain of
Penicillium oxalicum recovered from leaves of this plant in India [
364].
14. Miscellaneous Compounds
Resveratrol (
Figure 13) is a stilbene phytoalexin produced by many plants in response to biotic and abiotic injuries, and a reputed nutraceutical based on its antioxidant properties [
365]. A number of endophytic strains from
Vitis vinifera,
Vitis quinquangularis, and
P. cuspidatum belonging to the genera
Alternaria,
Aspergillus,
Botryosphaeria,
Cephalosporium,
Geotrichum,
Mucor and
Penicillium were found to be able to produce this compound; however, only a
Alternaria strain retained this capability after repeated culturing cycles [
366]. Another leaf strain of
Alternaria, together with root strains of
F. solani,
F. oxysporum and
F. proliferatum from
C. cajan, have been found to produce cajaninstilbene acid (
Figure 13), a related antioxidant compound originally characterized from the host plant [
367].
Figure 13.
Structures of miscellaneous compounds produced by endophytic fungi.
Figure 13.
Structures of miscellaneous compounds produced by endophytic fungi.
Mainly used as a food additive, curcumin (
Figure 13) is a diarylheptanoid compound of turmeric (
Curcuma longa) with potential pharmaceutical applications [
368], which has been recently found as a fermentation product of a previously-mentioned endophytic strain from
C. wenyujin [
39].
Falcarinol, also known as carotatoxin or panaxynol (
Figure 13), is a fatty alcohol reported from several unrelated plant species such as carrots (
Daucus carota) and other Apiaceae, red ginseng (
P. ginseng), ivies (
Hedera spp.) and other Araliaceae. It is considered a natural pesticide protecting roots from fungal diseases, and displayed some extent of activity against certain types of cancer [
369]. Production of this compound has been reported by a strain of
Paecilomyces sp. from
P. ginseng [
104].
Myrtucommulones are acylphloroglucinol compounds extracted from myrtle (
Myrtus communis) and related plants in the Myrtaceae considered as prospect pharmaceuticals based on consistent antibacterial, antimalarial, antioxidant, anti-inflammatory and antitumor properties. So far 13 analogues (myrtucommulones A-M) have been characterized from plant sources, and the production of myrtucommulone A (
Figure 13) and D has been recently reported from a myrtle endophytic strain of
Neofusicoccum australe (teleomorph
Botryosphaeria australis) [
370].
Finally, the widespread pentacyclic triterpenoid compound ursolic acid, also known as prunol or malol, and reported for its anticancer and cardioprotective properties [
371], has been recently found in extracts of a cited endophytic strain of
A. stygium [
51].
15. Future Perspectives
Data considered in this review highlight a continuously increasing number of plant-derived bioactive products also occurring as secondary metabolites of endophytic fungal strains. Therefore, it is quite easy to foresee that the finding of additional such compounds is under way and may disclose further applicative opportunities. As an example, the reported production of colchatetralene, a structural analog of colchicine, by an endophytic strain (
Aspergillus sp.) from seeds of
Gloriosa superba [
372] has paved the way for the possible production of this important drug by a microbial agent. Moreover, indications have been reported concerning the possible production of the secoiridoid glycoside gentiopicrin and of the cardiotonic drug digoxin by endophytic fungal strains, respectively from
Gentiana macrophylla [
373] and
Digitalis lanata [
374], and more circumstantiated studies are expected for these important compounds.
Within the above-considered categories, volatile compounds from essential oils particularly represent a mine to be dug more in depth. In fact benzophenone derivatives, which are biosinthetically related to xanthones and known as main components of the scent of species in a few botanical families such as Clusiaceae, Gentianaceae, Moraceae, Polygalaceae, Rosaceae and Thymelaceae, are also widespread secondary metabolites of fungi [
375]. Therefore, it would not be surprising if additional compounds previously reported from plants are also evidenced as fermentation products of endophytic strains [
376,
377,
378].
The likely finding by endophytic strains has been also anticipated for asperphenamate, a phenylalanine derivative recently attracting attention for its antitumor properties. In fact this compound was first characterized from
Aspergillus flavipes and a number of fungi which are also known for their endophytic habit. Therefore, its subsequently observed occurrence in many botanically unrelated plant species is considered to possibily derive by the biosynthetic ability of endophytic fungal strains [
379]. Besides the above-mentioned case of swainsonine, a more direct proof that some plant metabolites can be actually produced by their associated endophytic fungi has been provided by the finding of the mycotoxin alternariol, along with the related compounds altenusin and alternariol 5-
O-methyl ether, in foliar extracts of the medicinal plant
Polygonum senegalense harboring an
Alternaria strain [
380]. The latter compound had been also extracted from
Anthocleista djalonensis, a tree used in traditional medicine in West Africa [
381].
Further investigational opportunities arise from the ability by endophytic fungi to modify plant metabolites into novel bioactive compounds, and actually several examples of such a kind of biochemical interaction have been reported so far. A strain of
Xylaria sp. associated with
Cinchona pubescens was found to be able to convert the above-mentioned
Cinchona alkaloids into their 1-
N-oxides [
382]. Strains of
Diaporthe spp. from the tea plant (
Camellia sinensis) and from rhizome of
C. longa respectively performed the stereoselective oxidation of (+)-catechin and (–)-epicatechin into the corresponding 3,4-
cis-dihydroflavan derivatives [
383,
384], and the conversion of curcumin into a few colorless hydroderivatives [
385]. Moreover, an unidentified strain from yellow moonshed (
Archangelisia flava) has been reported to convert berberine into its 7-
N-oxide derivative [
386].
On the other end, endophytic fungi may also have a role in degradation of plant metabolites. As an example, a strain of
Phomopsis liquidambari from the stem of Chinese bishopwood (
Bischofia polycarpa) has been showed to degrade the phytoestrogen luteolin and additional phenolic compounds such as phenanthrene [
387], introducing the possible employment of endophytic fungal strains in bioremediation [
388].
However, there is no doubt that in the future the most intriguing research hint concerns the genetic bases of production of bioactive metabolites by these closely associated but phylogenetically unrelated organisms. Although a negationist point of view has been advanced concerning taxol biosynthesis by endophytes [
389], taken as a whole the over 300 cases mentioned in this review concerning homologous and heterologous endophytic strains producing plant bioactive compounds represent a compelling evidence that these micorganisms and their host plants interact at some level in the biosynthetic process. The main hypothesis is that these interactions may lead to horizontal gene transfers or genetic recombinations, from the plant to its endophytic counterpart and/or
vice-versa, originating novel specialized strains able to accumulate certain metabolites in the host tissues [
186], which would be a good reason to explain why in the end a mutualistic relationship is established. The fact that most of these compounds have been originally extracted from plants has generated a somehow misleading preconception that it is the endophytic microbe that for some reasons acquires the ability to produce a given plant metabolite. However, actually, the finding of taxol and other bioactive products from endophytic strains of plants which themselves are not reported as a source of the same compound could be considered as an indication that a reverse influence may be more likely. On the other hand, investigations concerning complex diterpenoid compounds, such as gibberellins and again taxol, have shown that the biosynthetic pathways in plant and endophytes may differ at some level [
5,
251], which is a clue for an independent development of the subtended genetic traits. However, in the end, the problem is again preconceptual, since it cannot be excluded that such a variation in the biosynthetic scheme may also exist within the categories of both plants and associated endophytes.
An even more complex situation could result if further evidence is provided that synthesis of bioactive metabolites in fungi is in turn influenced by other associated microbes. This is the case of rhizoxin which has been found to be actually synthesized by an endosymbiotic bacterium (
Burkholderia rhizoxinica) residing in the cytosol of the rice fungal pathogen
Rhizopus microsporus [
390,
391]. Moreover, such a kind of provision may well occur in endophytic fungi, considering that a widespread presence of endohyphal bacteria resulted in a survey on Ascomycetous endophytes of Cupressaceae [
392]. Moreover, a role of giant linear plasmids in the synthesis of antibiotic compounds in
Streptomyces has been proposed since over 25 years [
393]. If eventually confirmed for other metabolites, the role of these “third parties” could represent the means through which the biosynthetic abilities are interchanged between plants and their associated endophytes, and also explain why in a few cases this property is lost after repeated subculturing. This latter undesired aspect, which is reported to impair an efficient employment of endophytic fungi by the pharmaceutical industry, might also be consequential to more subtle events at the gene level, considering that the plant-endophyte interaction may involve promotion of gene transcriptions. Coherent with this possibility are the elicitating properties observed by endophytes on the synthesis of a number of bioactive plant products, with a few cases previously mentioned in this review [
193,
361,
362]. Finally, an intriguing elicitating function may also characterize other cohabiting endophytic strains not directly capable to produce a given metabolite, as shown in the case of strains of
Alternaria sp. and
Phomopsis sp. which consistently increased taxol synthesis by a taxol-producer
Paraconiothyrium sp. from
Taxus x
media in co-cultures [
394].
Quite clearly, the complex evolving scenario outlined in this paper is introductory to further advances in the elucidation of the genetic and biochemical bases of the synthesis of bioactive compounds, and the reflecting biocenotic interactions among plants, their associated endophytes and other involved microorganisms. New acquisitions in these fields will be fundamental in order to exploit microbial strains for a large-scale production of plant-derived drugs in controlled fermentative processes.