Deep-Sea Fungi Could Be the New Arsenal for Bioactive Molecules.

Growing microbial resistance to existing drugs and the search for new natural products of pharmaceutical importance have forced researchers to investigate unexplored environments, such as extreme ecosystems. The deep-sea (>1000 m below water surface) has a variety of extreme environments, such as deep-sea sediments, hydrothermal vents, and deep-sea cold region, which are considered to be new arsenals of natural products. Organisms living in the extreme environments of the deep-sea encounter harsh conditions, such as high salinity, extreme pH, absence of sun light, low temperature and oxygen, high hydrostatic pressure, and low availability of growth nutrients. The production of secondary metabolites is one of the strategies these organisms use to survive in such harsh conditions. Fungi growing in such extreme environments produce unique secondary metabolites for defense and communication, some of which also have clinical significance. Despite being the producer of many important bioactive molecules, deep-sea fungi have not been explored thoroughly. Here, we made a brief review of the structure, biological activity, and distribution of secondary metabolites produced by deep-sea fungi in the last five years.

1. Deep-Sea Fungi: A Novel Source of Bioactive Molecules

Antibiotics and antifungal drugs are the most commonly used drugs in the world, but their role in treating human diseases has been greatly reduced due to the development of pathogen resistance against these drugs. Scientists are now looking for new, untapped and renewable resources for the isolation of novel compounds to with clinical importance. Despite the fact that the ocean provides habitats to a huge number of microbes, both fungi and bacteria for thousands of years, the microbes of these extreme ecosystems and their potential for new drug discovery have not yet been fully realized due to methodological and technical limitations. Fungi are the most diverse and abundant eukaryotic organisms on the planet, and their presence in all possible extreme ecosystems make them an ideal source for investigations of new drug development. Scientists are interested in the extraction of novel and unique natural products, having clinical importance, from different organisms living in the extreme environments. In addition to terrestrial extreme environments, the ocean could also be considered a good reservoir of bioactive metabolites [1,2,3,4]. Fungi living in the deep-sea environments are known to produce novel bioactive compounds. Although, it is not fully understood why the fungi living in the extreme environments produce unique and novel products, it is assumed that fungal genome has evolved to make necessary adjustments in order to sustain life in such harsh conditions and might be involved in chemical defense and communication [5].

The ocean is considered to be one of the most diverse ecosystems. Compared to terrestrial and coastal ecosystems, the deep-sea (water depths below 1000 m) has a variety of extreme environments, such as temperatures ranging from 0 to 400 °C, lack of light and oxygen, high hydrostatic pressure up to 400 atm, and limited supply of nutrient substrates, making these habitats extremely difficult for life [6,7]. In order to inhabit such extreme ecosystems, organisms should have the potential to adjust to these conditions with different mechanism, such as regulating temperature, pH, and solute concentration, as well as the production of biomolecules to control DNA, protein, and lipid damage. This may be why microorganisms growing in these environments produce special metabolites.

Previously, drug investigators mainly considered bacteria, especially actinomycetes, as an important source of antifungal and antibacterial drugs. Cephalosporin C was the first compound derived from the marine fungus Cephalosporium sp. in 1949. After that, a number of important drugs— for instance, polyketide griseofulvin, terpenoid fusidic acid, cephalosporins, etc.—have been isolated from the marine fungi. Despite being the source of such important products, deep-sea fungi have not received full attention [8]. With the increasing demand for new drugs, scientists are now looking for new and unexplored resources for bioactive compounds, and the deep-sea consists of some extreme ecosystems that are worth exploring for new metabolites. Studies about isolating new bioactive molecules from marine environments are growing at an increasing rate, and hundreds of new compounds are reported every year; for instance, in 2017, a total of 448 new compounds were reported [9].

In this review, we present an overview of all those new and important bioactive metabolites isolated from deep-sea fungi during the last five years. We include only those molecules which were extracted from the deep-sea fungi associated with some kind of extreme environments, irrespective of its isolation from terrestrial counterparts, while all those compounds were excluded which were isolated from marine fungi and were not associated with extreme environments. This review will benefit all those who are interested in extreme-marine-environment fungi and their bioactive molecules. For more detailed information about other important secondary metabolites extracted from marine fungi, one should refer to our previous review papers [10,11,12].

2. Bioactive Compounds from Deep-Sea Fungi

According to the literature survey, we found 151 novel bioactive compounds isolated from marine fungi extracted from different extreme environments in the last five years. The majority of these compounds were isolated from two fungal genera i.e., Penicillium (63, 41.2% of the total compounds) and Aspergillus (43, 28.1% of the total compounds). Table 1 lists the detail of these compounds, which fall into different categories according to their structure.

Table 1

Secondary metabolites extracted from deep-sea fungi during 2013–2019.

Metabolites	Fungal Species	Source	Location	Depth (m) *	Bioactivity	Ref.
Polyketide
Methyl-isoverrucosidinol (1)	Penicillium sp. Y-50-10	Sulfur-rich Sediment	hydrothermal vent, Taiwan	_	Antibiotic	[13]
Penilactone A (2)	Penicillium crustosum PRB-2	Sediment	Prydz Bay, Antarctica	526	NF-kB inhibition	[14]
Aspiketolactonol (3)Aspilactonols A–F (4-9)Aspyronol (10)Epiaspinonediol (11)	Aspergillus sp. 16-02-1	Hydrothermal vent water	Lau Basin, Southwest Pacific Ocean,	2255	Cytotoxic	[15]
Ascomycotin A (12)Diorcinol (13)	Ascomycota sp. Ind19F07	Sediment	Indian Ocean	3614	Antibiotic	[16]
Engyodontiumones A–J (14-22)	Engyodontium album DFFSCS021	Sediment	South China Sea	3739	Cytotoxic	[18]
Lindgomycin (23)Ascosetin (24)	Lindgomycetaceae strains KF970 and LF327	Sediment	Greenland Sea, Baltic Sea	3650	Antibiotic	[17]
Nitrogen-containing compounds
Brevicompanines D–H (25-29)	Penicillium sp. F1	Sediment	_	5080	LPS-induced inflammation	[22]
Cyclopiamide B–J (30-38)	Penicillium commune DFFSCS026	Sediment	South China Sea	3563	Cytotoxic	[24]
Penipanoid A (39)Quinazolinone (40)	Penicillium paneum SD-44	Sediment	South China Sea	201	Cytotoxic	[23]
(±) Brevianamide R (41)	Aspergillusversicolor MF180151	Sediment	Bohai Sea, China	_	Antibacterial	[21]
Circumdatin F and G (42-43)	Aspergillus westerdijkiae SCSIO 05233	Sediment	South China Sea	4593	Cytotoxic	[20]
Oximoaspergillimide (44)Neohydroxyaspergillic (45)Neoaspergillic (46)	Aspergillus sp. (CF07002)	Water	Pacific Ocean off the coast of Panama		CytotoxicAntibiotic	[19]
Varioxepine A (47)	Paecilomyces variotii EN-291	Deep sea water	_	_	Antibiotic	[26]
Neoechinulin A (48)	Microsporum sp. (MFS-YL)	Red alga	Guryongpo, Korea	_	Cytotoxic	[25]
Polypeptide
Canescenin A and B (49-50)	Penicillium canescens SCSIO z053	Water	East China Sea	2013	Antibacterial	[27]
Clavatustide A and B (51-52)	Aspergillus clavatus C2WU	Hydrothermal vent crab	Taiwan Kueishantao	_	Cytotoxic	[29]
Aspergillamides C and D (53-54)Butyrolactone I (55)	Aspergillus terreus SCSIO 41008	Sponge	Guangdong, China	_	CytotoxicAntibiotic	[30]
Simplicilliumtides A–I (56-64)	Simplicillium obclavatum EIODSF 020	Sediment	East Indian Ocean	4571	Cytotoxic	[31]
Asperelines A–F (65-70)	Trichoderma asperellum	Sediment	Antarctic Penguin Island	_	Antibiotic	[28]
Esters
7-chlorofolipastatin (71)Folipostatin B (72)Unguinol (73)2-chlorounginol (74)2,7-dichlorounguinol (75)Nornidulin (76)	Aspergillus ungui NKH-007	Sediment	Suruga Bay, Japan	_	Anti-atheroscleroticCytotoxicAntibiotic	[32]
Phenolic
Pestalotionol (77)	Penicillium sp. Y-5-2	Hydrothermal vent water	Kueishantao off Taiwan	_	Antibiotic	[33]
Aspergilol G–I (78-80)Coccoquinone A (81)	Aspergillus versicolor SCSIO 41502	Sediment	South China Sea	2326	Anti-HSV-1AntioxidantAntifouling	[34]
Piperazine
Fusaperazine F (82)	Penicillium crustosum HDN153086	Sediment	Prydz Bay, Antarctica	_	Cytotoxic	[35]
N-methyl-pretrichodermamide B (83)Pretrichodermamide C (84)	Penicillium sp. (WN-11-1-3-1-2)	Hypersaline sediment	Wadi El-Natrun, Egypt	_	Cytotoxic	[36]
(±) 7,8-epoxy-brevianamide Q (85)(±) 8-hydroxy-brevianamide R (86)(±) 8-epihydroxy-brevianamide R (87)Brevianamide R (88)Versicolorin B (89)	Aspergillus versicolor MF180151	Sediment	Bohai Sea, China	_	Antibiotic	[21]
Dichotocejpins A (90)6-deoxy-5a,6-didehydrogliotoxin (91)Gliotoxin (92)Acetylgliotoxin (93)6-acetylbis(methylthio)-gliotoxin (94)1,2,3,4-tetrahydro-2-methyl-3-methylene-1,4-dioxopyrazino [1,2-a] indole (95)	Dichotomomyces cejpii FS110	Sediment	South China Sea	3941	α-Glucosidase inhibitionCytotoxic	[37]
Terpenoid
Brevione F–I (96-99)	Penicillium sp. (MCCC 3A00005)	Sediment	Pacific Ocean	5115	CytotoxicHIV-1 inhibition	[22,38]
Dehydroaustin (100)Dehydroaustinol (101)7-hydroxydehydroaustin (102)Austinone (103)Austinol (104)Austin (105)Austinolide (106)	Penicillium sp. Y-5-2	Hydrothermal vent water	Kueishantao off Taiwan	8	Antibacterial Anti-insectal	[33]
1-chloro-3β-acetoxy-7-hydroxytrinoreremophil-1,6,9-trien-8-one (107)Eremophilane-type sesquiterpenes (108)Eremofortine C (109)	Penicillium sp. PR19N-1	Sediment	Prydz Bay, Antarctica	526	Cytotoxic	[40,41]
Guignarderemophilane F (110)	Penicillium sp. S-1-18	Sediment	Antarctic	1393	Antibacterial	[42]
Spirograterpene A (111)Conidiogenone C and I (112-113)	Penicillium granulatum MCCC 3A00475	Water	Prydz Bay of Antarctica	2284	Antiallergic	[46]
Aspewentin A and D–H (114-118)Asperethers A–E (121-125)Asperolides D and E (119-120)	Aspergillus wentii SD-310	Sediment	South China Sea	2038	AntimicrobialCytotoxicAnti-inflammatory	[39,43,44]
(7S)-(+)7-O-methylsydonol (126)(7S,11S)-(+)-12-hydroxysydonic acid (127)7-deoxy-7,14-didehydrosydonol (128)(S)-(+)-sydonol (129)	Aspergillus sydowii	Sediment	Hsinchu, Taiwan	_	Anti-inflammatory	[47]
6b,9a-dihydroxy-14-p-nitrobenzoylcinnamolide (130)Insulicolide A (131)	Aspergillus ochraceus Jcma1F17	Marine alga Coelarthrum sp.	South China Sea	_	AntiviralCytotoxic	[45]
Other compounds
Sterolic acid (132)	Penicillium sp. MCCC 3A00005	Sediment	East Pacific Ocean	5115	Cytotoxic	[38]
Dicitrinone B (133)	Penicillium citrinum	Sediment	Langqi Island, Fujian, China	_	Antitumor	[49]
Penipacids A–F (134-139)	Penicillium paneum SD-44	Sediment	South China Sea	_	Cytotoxic	[48]
Butanolide A (140)	Penicillium sp. S-1-18	Sediment	Antarctic seabed	1393	Cytotoxic	[42]
7-Methoxycyclopeptin (141)7-Methoxy dehydro cyclopeptin (142)7-Methoxy cyclopenin (143)9-Hydroxy-3-methoxyviridicatin (144)	Aspergillus versicolor XZ-4	Hydrothermal vent crab	Kueishantao, Taiwan		Antibiotic	[50]
Wentilactone A and B (145-146)	Aspergillus dimorphicus SD317	Sediment	South China Sea	2038	Antitumor	[51]
Penicillisocoumarin A–D (147-150)Aspergillumarins B (151)	Penicillium sp. Y-5-2	Hydrothermal vent water	Kueishantao off Taiwan	8	Antibacterial	[33]

* Depth represents water depth below the surface.

2.1. Polyketide Compounds

Twenty-four polyketide compounds (1–24; Figure 1) with important biological activities were isolated from fungi extracted from different deep-sea environments. Among them, compounds 1 and 2 were isolated from Penicillium spp., which showed antibiotic activity (MIC of 32 μg/mL against Bacillus subtilis) and nuclear factor NF-kB inhibition activity, respectively [13,14]. Compounds 3–11 were from Aspergillus sp. 16-02-1, which exhibited cytotoxicity (with a 10%–80% inhibition rate at 100 μg/mL against various cancer cell lines i.e., K562, HL-60, HeLa, and BGC-823) [15]. Similarly, compounds 12–24 were isolated from the species belonging to Ascomycetes, Engyodontium, and Lindgomycetaceae, out of which compounds 12–13 and 23–24 showed strong antibiotic activities against Bacillus subtilis, Acinetobacter baumannii, Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, Staphylococcus epidermidis, and Propionibacterium acnes, while compounds 14–22 exhibited strong cytotoxic activity (IC50 4.9 µM) against U937 cells (Table 1) [16,17,18].

Figure 1

Structures of polyketide secondary metabolites obtained from deep-sea fungi.

2.2. Nitrogen-Containing Compounds

Twenty-four novel alkaloid-bioactive compounds (25–48; Figure 2) have been reported from deep-sea fungi since 2013, out of which compounds 25–40 were isolated from Penicillium spp., and showed cytotoxic activities against BV2 cell (IC50 of 27–45 µg/mL), brine shrimp (IC50 of 14.1 to 38.5 µg/mL), SMMC-7721 (IC50 of 54.2 µM), BEL-7402 ((IC50 of 17.5 µM), and BEL-7402 (IC50 of 19.8 µM) [19,20,21]. Compounds 41–46 were identified from Aspergillus spp., in which compounds 41 and 45–46 displayed antibiotic activity (MIC of 30 to 40 µg/mL) against BCG, Candida albicans, Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus cereus, Klebsiella pneumoniae, and Escherichia coli, while compounds 47 and 48 were extracted from other genera and showed antimicrobial activity (MIC between 16 and 64 µg/mL against Escherichia coli, Aeromonas hydrophila, Micrococcus luteus, Staphylococcus aureus, Vibrio anguillarum, Vibrio harveyi, and Vibrio parahaemolyticus) and cytotoxic activity against human cervical carcinoma HeLa, respectively [22,23,24,25,26].

Figure 2

Bioactive alkaloid compounds isolated from deep-sea fungi.

2.3. Polypeptides

Twenty-two polypeptides with novel structures (49–70; Figure 3) were reported from fungi inhabiting different marine environments during 2013–2019. Compounds 49 and 50 were isolated from Penicillium canescens and displayed antibiotic activity against Bacillus amyloliquefaciens and Pseudomonas aeruginosa at 100 µM, while compounds 51–55 were extracted from Aspergillus spp., in which 51–54 showed cytotoxic activity (IC50 of 15–25 μg/mL) against HepG2, SMMC-7721, Bel-7402, and human glioma U87 cell lines, while compound 55 showed inhibitory effects (IC50 value of 5.11 μmol/L) against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) [27,28,29,30]. However, compounds 56–64, which were obtained from Simplicillium obclavatum, and 65–70, obtained from Trichoderma asperellum, displayed cytotoxicity (IC50 of 39.4–100 µM) against human leukemia HL-60 and K562 cell lines and antibiotic activity (IC50 of 39.4–100 µM) against Gram-positive bacteria (e.g., Bacillus amyloliquefaciens, Staphylococcus aureus) and Gram-negative bacteria (e.g., Pseudomonas aeruginosa and Escherichia coli), respectively [28,31].

Figure 3

Bioactive polypeptides isolated from deep-sea fungi.

2.4. Ester and Phenolic Derivatives

Six new ester derivatives (71–76; Figure 4) were extracted from Aspergillus ungui NKH-007 and showed inhibition of sterol O-acyltransferase (SOAT) enzymes in Chinese hamster ovary (CHO) cells and are thus considered to be good candidates for an anti-atherosclerotic agent [32]. Five new phenolic compounds (77–81; Figure 4) isolated from Penicillium sp. and Aspergillus versicolor showed potent activity against Staphylococcus aureus and Bacillus subtilis, with MIC values of 2–8 μg/mL [33,34]. However, compounds 78–81 expressed antiviral activity toward HSV-1, with EC50 values of 3.12–6.25 μM [34].

Figure 4

Ester and phenolic derivatives obtained from deep-sea fungi.

2.5. Piperazine Derivatives

Fourteen new piperazine derivatives (82–95; Figure 5) reported from marine fungi during the last five years. These derivatives were isolated from genera of Penicillium, Aspergillus, and Dichotomomyces collected from deep-sea sediments. Compounds 82–84 showed strong cytotoxicity with IC50 of 1.7 and 2 µM against K562 and mouse lymphoma cell line, respectively; similarly, compounds 91–95 also showed strong cytotoxic activity [35,36,37]. Compounds 85–89 showed antibacterial activity against Staphylococcus aureus with the MIC values of 6.25–12.5 µg/mL [21]. The new compound 90 also showed stronger inhibition activity against α-glucosidase with IC50 value of 138 µM [37].

Figure 5

Piperazine derivatives isolated from deep-sea fungi.

2.6. Terpenoid Compounds

Thirty-six new and important bioactive terpenoids (96–131; Figure 6) have been isolated from marine fungi extracted from the deep-sea sediments since 2013. Compounds 96–113 were isolated from Penicillium spp., while compounds 114–131 were extracted from Aspergillus spp. Breviones (96–99), isolated from the deepest sediment-derived fungus Penicillium sp. (5115 m depth), displayed diverse activities, such as cytotoxicity against HeLa, MCF-7, and A549 cells with IC50 values of 7.44 to 32.5 µM, respectively, and growth inhibition of HIV-1 with EC50 value of 14.7 µM against C8166 cells [22,38]. Compounds 100–110 showed antibiotic and inhibition activities against silkworm, while 20-nor-isopimarane diterpenoids, including aspewentins (114–118), asperethers (121–125), asperoloids (119–120), and compounds 130 and 131, showed cytotoxic activities [33,39,40,41,42,43,44,45]. However, the spirocyclic diterpenes (111–113) exhibited strong anti-allergic effect with 18% inhibition at 20 μg/mL [46]. Interestingly, four new compounds (126–129) were extracted from hydrothermal vent-derived Aspergillus sydowii, through activation of a new pathway for secondary metabolite production by the addition of a 5-azacytidine (a DNA methyltransferase inhibitor). These compounds showed anti-inflammatory and antidiabetic activities and are thus the first secondary metabolites isolated from fungi which have both antidiabetic and anti-inflammatory activities [47].

Figure 6

Structures of terpenoid secondary metabolites obtained from deep-sea fungi.

2.7. Other Unrelated Compounds

Twenty secondary metabolites with different structures were isolated from deep-sea fungi, mainly from Penicillium spp. and Aspergillus spp. (132–151; Figure 7). Penipacids A–F (134–139), polyoxygenated sterol (132), dicitrinone B (133) and butanolide A (140), which were isolated from deep-sea sediments-derived Penicillium spp., showed cytotoxic activities against RKO, MCF-7, PTP1B and A375 cancer cell lines with IC50 values of 8.4–28.4 µM [38,42,48,49]. Similarly, four isocoumarins, penicillisocoumarin A–D (147–150), and an isocoumarins aspergillumarin B (151) were also isolated from Penicillium which showed weak antibacterial activities [33]. Four antibiotic cyclopenin derivatives compounds (141–144) and a series of antitumor wentilactones (145,146) were isolated from Aspergillus spp. [50,51].

Figure 7

Bioactive metabolites derived from deep-sea fungi.

3. Conclusions and Perspective

The results of current studies indicate that the deep-sea extreme environmental fungi are one of the rich and unexploited sources of important medicinal lead compounds. Most of the fungi (e.g., Penicillium spp. and Aspergillus spp.) living in the extreme environments of the deep-sea have the potential to synthesize new bioactive compounds. However, the research on deep-sea fungi and their metabolites is very limited due to the difficulty of sampling and the limitation of culture technology. Thanks to the advances in genome technology and the implementation of the deep-sea drilling program, novel compounds with great biological activities are expected from these fungi in the near future. From the literature review, we can say these fungi from the extreme environments have the potential to produce clinically important natural products. The compounds we discussed in this review show strong bioactivities and might have the potential to be a future anticancer drug. Among them, terpenoid derivatives were the most important and abundant compound category which were mainly isolated from deep-sea derived Penicillium spp. and Aspergillus spp. This class of compounds showed strongest antibiotic and cytotoxic activities as compared to other classes of compounds and has the potential to be a future candidate for anticancer drugs, especially brevione, which was isolated from the deepest part of the sea and showed the strongest cytotoxic activity.