Skip to main content
NCBI home page
Mycobiology logoLink to Mycobiology
. 2025 Nov 3;53(6):794–806. doi: 10.1080/12298093.2025.2578051

Endophytic Fungi Isolated from the Brown Alga Sargassum thunbergii in Coastal Korea

Ji-Won Kim 1,#, Yu-Na Choi 1,#, Eun-Ju Kim 1, Ahn-Heum Eom 1,✉
PMCID: PMC12584843  PMID: 41195061

Abstract

Endophytic fungi associated with seaweeds have increasingly attracted attention due to their ecological roles and potential as biological resources. In this study, fungal endophytes were isolated from the brown alga Sargassum thunbergii, collected in intertidal habitats of Seongsan-eup, Jeju-do and Buan-gun, Jeollabuk-do, Korea. Identification was carried out using an integrative approach that combined morphological characterization with multilocus phylogenetic analyses of ITS, LSU, SSU, TUB2, RPB2, and CaM sequences. Six fungal taxa were identified as new records for Korea: Botryotrichum murorum, Hypomontagnella spongiphila, Subramaniula cristata, Parachaetomium perlucidum, Palmiascoma acericola, and Aspergillus clavatophorus. Detailed morphological descriptions and phylogenetic analysis supporting the identification of these species are provided. These findings expand our knowledge of fungal diversity associated with S. thunbergii and highlights the ecological importance and potential applications of seaweed-associated endophytes in Korea.

Keywords: Endophytic fungi, Sargassum thunbergii, phylogenetic analysis, seaweed-associated fungi

1. Introduction

Sargassum thunbergii (Mertens ex Roth) Kuntze is a perennial brown alga endemic to the northwestern Pacific, forming extensive populations along the rocky intertidal zones of Korea, China, and Japan [1,2]. These dense assemblages provide essential habitats, refuges, and nursery grounds for diverse marine organisms [1,3]. In addition, they contribute to nutrient cycling and coastal ecosystem stability by accumulating heavy metals such as zinc, cadmium, copper, and arsenic, and by assimilating nitrogen and phosphorus from eutrophic seawater [2,4,5]. Due to its high productivity and strong tolerance to environmental stressors such as heat, light, and desiccation, S. thunbergii play an ecologically important role in Northeast Asian coastal ecosystems and has recently been recognized as a promising candidate for seaweed forest restoration and aquaculture [1,2].

Endophytic fungi are microorganisms that reside within host tissues without causing pathological symptoms, and they are distributed across both terrestrial and marine ecosystems [6–11]. Marine endophytes associated with seaweeds are predominantly ascomycetous fungi, including genera such as Aspergillus, Penicillium, Fusarium, Chaetomium, Alternaria, and Cladosporium, many of which are also commonly reported as endophytes of terrestrial plants [8,12–14]. The distinctive conditions of marine environments, including high salinity, temperature fluctuations, and elevated hydrostatic pressure, strongly influence their diversity and metabolic activity [15–17]. These stressors are known to induce the expression of unique gene clusters or lead to variations in secondary metabolite production [6,18]. For example, certain marine fungi exhibit optimal growth only within specific salinity and temperature ranges, suggesting distinct biochemical pathways compared to their terrestrial counterparts [17–19].

The ecological importance of endophytic fungi associated with marine macroalgae is evident in their role as chemical mediators within host–environment interactions [6,9]. By producing diverse secondary metabolites, these fungi protect their hosts against pathogens, grazers, ultraviolet radiation, and fluctuations in salinity, thereby shaping complex ecological relationships in marine communities [14,20]. These bioactive compounds, including those with anti­microbial, antifungal, antioxidant, anti-inflammatory, and anticancer activities, also provide a valuable resource for human use [8,9]. Accordingly, marine endophytic fungi are increasingly recognized as promising sources of novel secondary metabolites with potential applications in pharmaceuticals, aquaculture, agriculture, and environmental biotechnology, while their biodiversity and functional capacities remain underexplored [9,16,18,19,21].

Globally, studies on algicolous fungi have advanced considerably, leading to the discovery of novel taxa and their metabolites [6]. In Korea, however, systematic investigations of fungal diversity associated with marine macroalgae remain limited, and only recently have unrecorded species and their potential ecological significance been reported [10,11,22]. Nevertheless, the diversity and ecological roles of endophytic fungi in Korean seaweeds remain poorly understood compared to those of terrestrial endophytes. To address this knowledge gap, we conducted morphological examinations and multilocus phylogenetic analyses of endophytic fungi isolated from the brown alga Sargassum thunbergii collected in Seongsan-eup, Jeju-do and Buan-gun, Jeollabuk-do. As a result, six fungal species are reported as new record for Korea - Botryotrichum murorum, Hypomontagnella spongiphila, Subramaniula cristata, Parachaetomium perlucidum, Palmiascoma acericola, and Aspergillus clavatophorus - thereby contributing to a deeper understanding of endophytic fungal diversity in Korean marine ecosystems.

2. Materials and methods

2.1. Sample collection and preparation

From August 2024 to April 2025, samples of Sargassum thunbergii were collected from intertidal zones of Seongsan-eup (33°27’36.87ʺ N 126°56’5.15ʺ E, 33°24′07.3″ N 126°54′20.4″ E), Jeju-do and Buan-gun (35°38′12.44″ N 126°27′42.10″ E, 35°37′6.0″ N 126°28′0.12″ E), Jeollabuk-do, Korea. Only healthy thalli without visible wounds or disease symptoms were selected. Samples were placed in clean plastic bags containing seawater and transported to the laboratory within 12 h.

2.2. Surface sterilization and fungal isolation

To remove surface contaminants, algal thalli were first rinsed with tap water [23] and then cut into small fragments measuring approximately 1.5 × 0.5 cm. The fragments were surface-sterilized by immersion in 70% ethanol for 15 s, followed by a rinse with sterile seawater to eliminate residual ethanol [24]. After sterilization, the samples were gently blotted dry with sterile cotton [25] and inoculated onto two types of media: Dichloran Rose Bengal Chloramphenicol (DRBC; MBcell, Seoul, Korea) prepared with seawater and Potato Dextrose Agar (PDA; Difco Lab., Detroit, USA) supplemented with chloramphenicol [23]. All plates were incubated at 25 °C in the dark, and emerging fungal hyphae were subcultured onto fresh PDA to establish pure isolates [26].

2.3. Culturing and morphological characterization

Fungal isolates were subsequently cultured on malt extract agar (MEA; Kisan Bio, Seoul, Korea) and PDA at 25 °C in the dark. Colony characteristics, including pigmentation, surface texture, and margin features, were documented through visual observation [12]. Reproductive structures were examined using a stereomicroscope (SZX7, Olympus, Tokyo, Japan), and spores examined using a light microscope (Axio Imager A2, Carl Zeiss, Oberkochen, Germany), to document diagnostic morphological features.

2.4. DNA extraction, PCR and sequencing

Genomic DNA was extracted from cultured mycelia using the HiGene Genomic DNA Prep Kit (Biofact, Daejeon, Korea). Polymerase chain reaction (PCR) was carried out to amplify internal transcribed spacer (ITS) region [27], partial sequences of additional loci commonly employed for fungal identification, including the large subunit ribosomal RNA (LSU) [28,29], β-tubulin (TUB2) [30–32], RNA polymerase II second largest subunit (RPB2) [33], the small subunit ribosomal RNA (SSU) [34], and calmodulin (CaM) [35]. Universal primers were used for each locus, and the corresponding primer sets and PCR conditions are summarized in Table 1. Sequencing was performed by Macrogen Co., Ltd. (Sejong, Korea), and the resulting sequences were compared with entries in the NCBI database using the Basic Local Alignment Search Tool (BLAST) to confirm taxonomic identity.

Table 1.

DNA region, primer sets and PCR condition for molecular phylogenetic analysis.

DNA region Primer pair PCR condition Fungal strain
ITS ITS1F/ITS4 [27] 95 °C: 20 s, 50 °C: 40 s,a
72 °C: 60 s (35 cycles)d 		KNUE 24S467, KNUE 24S533, KNUE 25S011, KNUE 25S025, KNUE 25S034, KNUE 25S318
LSU LR0R/LR5 [28] 95 °C: 20 s, 44 °C: 40 s,a
72 °C: 60 s (35 cycles)d 		KNUE 24S467, KNUE 24S533, KNUE 25S025, KNUE 25S034
  NL1/NL4 [29] 94 °C: 45s, 60 °C 45 s (touchdown)*,c
72 °C 120 s (45 cycles)d 		KNUE 25S011
TUB2 Bt2a/Bt2b [30] 95 °C: 20 s, 55 °C: 40 s,a
72 °C: 60 s (35 cycles)d 		KNUE 25S318
  T1/Btub4Rd [31,32] 94 °C: 60 s, 55 °C: 60 s,b
72 °C: 60 s (30 cycles)e 		KNUE 24S467, KNUE 25S011
KNUE 25S025
RPB2 RPB2-5F2/RPB2-7cR [33] 95 °C: 45 s, 56 °C: 80 s,b
72 °C: 120 s (35 cycles)e 		KNUE 24S467, KNUE 25S011
KNUE 25S025, KNUE 25S034
KNUE 25S318
SSU NS1/NS4 [34] 94 °C: 45 s, 55 °C: 45 s,c
72 °C: 60 s (35 cycles)e 		KNUE 25S034
CaM cmd5/cmd6 [35] 95 °C: 20 s, 55 °C: 40 s,a
72 °C: 60 s (35 cycles)d 		KNUE 25S318
Open in a new tab

aInitiation step of 95 °C: 2 min.

bInitiation step of 95 °C: 5 min.

cInitiation step of 94 °C: 4 min.

dFinal elongation step of 72 °C: 5 min and final hold at 4 °C.

eFinal elongation step of 72 °C: 10 min and final hold at 4 °C.

*Touchdown PCR: annealing temperature decreased stepwise from 60 °C (5 cycles) to 58 °C (5 cycles), and then fixed 56 °C (35 cycles).

2.5. Phylogenetic analysis

Phylogenetic trees were inferred using Maximum Likelihood (ML) method implemented in MEGA11 [36]. The best-fit substitution models were determined by model selection tests based on the lowest Bayesian Information Criterion (BIC) value and applied as follows: K2 + G (KNUE 25S034), K2 + G + I (KNUE 24S533), T92 + G (KNUE 24S467, KNUE 25S011 and KNUE 25S025), and TN93 + G (KNUE 25S318). The robustness of the tree topology was evaluated using bootstrap analysis with 1000 replicates. All fungal isolates were deposited in the National Institute of Biological Resources (NIBR), and their sequence data were submitted to NCBI GenBank.

3. Results

3.1. Phylogenetic analysis

Both locus-specific BLAST searches and multilocus phylogenetic analyses supported the molecular identification of the six isolates. BLAST comparisons of ITS, LSU, SSU, TUB2, CaM, and RPB2 sequences consistently showed high similarity to their respective reference strains, as summarized in Table 2. Phylogenetic trees based on concatenated datasets further confirmed the placement of the isolates within well-supported clades corresponding to Botryotrichum murorum, Subramaniula cristata, Parachaetomium perlucidum, Hypomontagnella spongiphila, Palmiascoma acericola, and Aspergillus clavatophorus (Figures 1–6).

Table 2.

Sequence similarities of the six fungal isolates with reference strains based on BLAST searches. T indicates an ex-type culture.

Strain ID Taxon
(Identified species)		 Locus Reference strain Identity (%) Accession No.
KNUE 24S467 Botryotrichum murorum ITS CBS 163.52 100.00 KX976591
LSU CBS 163.52 99.81 KX976716
TUB2 CBS 163.52 100.00 KX976933
RPB2 CBS 163.52 99.70 KX976815
KNUE 25S011 Subramaniula cristata ITS CBS 156.52 T 100.00 KX976690
LSU CBS 156.52 T 99.81 KX976788
TUB2 CBS 156.52 T 99.85 KX977038
RPB2 CBS 156.52 T 99.80 KX976903
KNUE 25S025 Parachaetomium perlucidum ITS CBS 141.58 T 99.62 MK919308
TUB2 CBS 141.58 T 99.09 MK919422
RPB2 CBS 141.58 T 100.00 MK919365
KNUE 24S533 Hypomontagnella spongiphila ITS CLL 205 T 98.76 MK131719
LSU CLL 205 T 99.30 MK131717
RPB2 CLL 205 T 98.40 MK135890
TUB2 CLL 205 T 94.07 MK135892
KNUE 25S034 Palmiascoma acericola ITS CGMCC 3.24963 T 99.43 OR253099
LSU CGMCC 3.24963 T 99.29 OR253251
SSU CGMCC 3.24963 T 99.61 OR253178
RPB2 CGMCC 3.24963 T 98.73 OR262121
KNUE 25S318 Aspergillus clavatophorus ITS NRRL 25874 T 100.00 KY087772
TUB2 NRRL 25874 T 100.00 KY117836
CaM NRRL 25874 T 98.84 KY068323
RPB2 NRRL 25874 T 99.88 KY118014
Open in a new tab

Figure 1.

Figure 1.

Open in a new tab

Maximum likelihood tree of Botryotrichum murorum KNUE 24S467 based on concatenated sequences of internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU), RNA polymerase II second largest subunit (RPB2), and β-tubulin (TUB2) DNA sequences. Subramaniula cristata was used as an outgroup. Bootstrap values greater than 50% (1,000 replicates) are shown at the nodes. T indicates an ex-type culture.

Figure 2.

Figure 2.

Open in a new tab

Maximum likelihood tree of Subramaniula cristata KNUE 25S011. The tree is based on concatenated sequences of internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU), β-tubulin (TUB2) and RNA polymerase II second largest subunit (RPB2) DNA sequences. Humicola fuscoatra was used as an outgroup. Bootstrap values greater than 50% (1,000 replicates) are shown at the nodes. T indicates an ex-type culture.

Figure 3.

Figure 3.

Open in a new tab

Maximum likelihood tree of Parachaetomium perlucidum KNUE 25S025. The tree is based on concatenated sequences of internal transcribed spacer (ITS), β-tubulin (TUB2) and RNA polymerase II second largest subunit (RPB2) DNA sequences. Chaetomium globosum was used as an outgroup. Bootstrap values greater than 50% (1,000 replicates) are shown at the nodes. T indicates an ex-type culture.

Figure 4.

Figure 4.

Open in a new tab

Maximum likelihood tree of Hypomontagnella spongiphila strain KNUE 24S533, based on concatenated sequences of the internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU), the second largest subunit of RNA polymerase II (RPB2), and β-tubulin (TUB2) DNA sequences. Xylaria hypoxylon was used as the outgroup. Bootstrap values greater than 50% (1,000 replicates) are shown at the nodes. T indicates an ex-type culture.

Figure 5.

Figure 5.

Open in a new tab

Maximum likelihood tree of Palmiascoma acericola KNUE 25S034. The tree is based on concatenated sequences of internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU), small subunit ribosomal RNA (SSU) and RNA polymerase II second largest subunit (RPB2) DNA sequences. Murilentithecium clematidis was used as an outgroup. Bootstrap values greater than 50% (1,000 replicates) are shown at the nodes. T indicates an ex-type culture.

Figure 6.

Figure 6.

Open in a new tab

Maximum likelihood tree of Aspergillus clavatophorus strain KNUE 25S318, based on concatenated sequences of the internal transcribed spacer (ITS), β-tubulin (TUB2), calmodulin (CaM), and second largest subunit of RNA polymerase II (RPB2). Hamigera avellanea was used as the outgroup. Bootstrap values greater than 50% (1,000 replicates) are shown at the nodes. T indicates an ex-type culture.

3.2. Taxonomy

Botryotrichum murorum (Corda) X. Wei Wang & Samson, Stud. Mycol. 84: 164 (2016) [MB#818837] (Figure 7)

Figure 7.

Figure 7.

Open in a new tab

Morphology of Botryotrichum murorum KNUE 24S467. A-B Colonies after 7 days of growth at 25 °C. The left side shows the front view and the right side shows the reverse view, on malt extract agar (A), and potato dextrose agar (B). Ascomata observed on the colony surface on PDA (C) and on a slide glass (D). Asci with immature ascospores were observed (E), and mature and immature ascospores were observed (F). Scale bars: C = 500 μm; D = 200 μm; E, F = 20 μm.

Morphological characteristics: After 7 days of incubation at 25 °C in the dark, colonies on MEA reached 22.0–27.0 mm in diameter. Colonies were pale yellowish pink, irregular in outline, relatively even and cottony in texture; the reverse was dark purplish gray at the center, fading to pale yellowish pink toward the margin. On PDA, colonies reached 27.0–30.0 mm in diameter. Colonies were yellowish gray, nearly circular in outline, uneven and distinctly lined with fine striations, cottony in texture; the reverse was wrinkled, grayish olive at the center, becoming progressively lighter toward the periphery. Ascomata were formed as perithecia, superficial, ostiolate, dark brown to black; wall brown. Ascomatal hairs were flexuous to undulate, typically more than twice the length of the ascoma. Asci were fasciculate, fusiform to occasionally clavate; spore-bearing portion (27.1–)36.4(–42.3) × (17.0–)18.9(–24.3) μm (n = 6), containing eight irregularly arranged ascospores. Ascospores were olivaceous-brown at maturity, ellipsoidal to fusiform, (13.0–)15.6(–14.7) × (7.8–)8.3(–8.8) μm (n = 20).

Specimen examined: Korea, Jeju-do, Seongsan-eup, 33°27′36.868″ N 126°56′5.136″ E, 1 Aug. 2024, isolated from S. thunbergii, strain KNUE 24S467 (NIBRFGC000513996); GenBank: PX149079 (ITS), PX149082 (LSU), PX207095 (RPB2), PX204774 (TUB2).

Notes: B. murorum was originally described as Chaetomium murorum Corda in 1837 and was transferred to Botryotrichum in 2016 [37]. Reference strains DTO 324-G9 and DTO 333-E6 are characterized by olivaceous-brown, ellipsoidal to fusiform ascospores with an apical germ pore, (12–)12.5–15(–16.5) × (7–)7.5–8.5 μm, and fusiform asci with spore-bearing portions 27–45 × 12.5–19 μm [37]. The morphology of strain KNUE 24S467 was consistent with these descriptions. Although the two reference strains exhibited slight morphological variation, phylogenetic analyses revealed no sequence divergence, and these differences were interpreted as intraspecific morphological diversity within B. murorum [37].

Subramaniula cristata (L.M. Ames) X. Wei Wang & Samson, Stud. Mycol. 84: 212 (2016) [MB#818853] (Figure 8)

Figure 8.

Figure 8.

Open in a new tab

Morphology of Subramaniula cristata KNUE 25S011. A-B Colonies after 7 days of growth at 25 °C are shown. The left side displays the front view and the right side displays the reverse view, observed on malt extract agar (A) and potato dextrose agar (B), respectively. Ascomata were observed on the colony surface on PDA (C) and a slide glass (D). Asci with immature ascospores were observed (E), and mature ascospores were observed (F). Scale bars: C = 500 μm; D = 200 μm; E–F = 20 μm.

Morphological characteristics: After 7 days of incubation at 25 °C in the dark, colonies on MEA reached 20.0–22.0 mm in diameter. Colonies were circular with a dense hyphal mat, flat elevation, and smooth surface; the colony surface was dark brown, with a distinct lighter-colored ring at the margin, and the reverse exhibited a similar coloration and overall morphology. On PDA, colonies reached 27.0–28.0 mm in diameter. Colonies were circular, densely compacted with fine, irregular wrinkles, and exhibited a flat profile with a smooth surface. The margin was yellowish-gray, gradually transitioning to a pale yellowish brown toward the center; the reverse closely resembled the surface in both color and structure. Ascomata were superficial, ostiolate, with brown walls and ascomatal hairs. Asci were clavate, eight-spored; spore-bearing portion (25.8–)29.1(–34.6) × (11.6–)12.2(–12.9) μm (n = 6). Ascospores were olive-brown at maturity, ovate to coffee bean-shaped with a central depression, (8.4–)10.5(–12.5) × (4.5–)5.4(–6.2) μm (n = 20).

Specimen examined: Korea, Jeollabuk-do, Buan-gun, 35°37′6.0″ N 126°28′0.12″ E, 21 Mar. 2025, isolated from S. thunbergii, strain KNUE 25S011 (NIBRFGC000513998); GenBank: PX121655 (ITS), PX121657 (LSU), PX127654 (TUB2), PX127655 (RPB2).

Notes: S. cristata was originally described as Chaetomium cristatum L.M. Ames in 1950, discovered on a paper carton under test at the Tropical Testing Chamber, Fort Belvoir, Virginia [38]. It was transferred to Subramaniula in 2016 [37]. The morphological features of strain KNUE 25S011 correspond well with the protologue, which described clavate asci (23–40 × 9–12 μm) and olive-brown, ovate to umbonate ascospores (9–11 × 5–7 μm) [37].

Parachaetomium perlucidum (Sergeeva) X. Wei Wang & Houbraken, Stud. Mycol. 101: 189 (2022) [MB#830930] (Figure 9)

Figure 9.

Figure 9.

Open in a new tab

Morphology of Parachaetomium perlucidum KNUE 25S025. A-B Colonies after 7 days of growth at 25 °C are shown. The left side displays the front view and the right side displays the reverse view, observed on malt extract agar (A) and potato dextrose agar (B), respectively. Ascoma were observed on the colony surface on PDA (C) and on a slide glass (D). Asci with immature ascospores were observed (E), and mature ascospores were observed (F). Scale bars: C = 200 μm; D = 100 μm; E = 20 μm; F = 10 μm.

Morphological characteristics: After 7 days of incubation at 25 °C in the dark, colonies on MEA reached 45.0–47.0 mm in diameter. Colonies were circular with densely interwoven hyphae, flat in elevation, and with a smooth, cottony surface; the colony surface was white at the center, becoming yellowish toward the margin, while the reverse was pale yellowish white with faint yellow pigmentation. On PDA, colonies reached 28.0–40.0 mm in diameter. Colonies were circular, compact, densely packed, flat in elevation, and with a smooth, cottony texture; the surface was entirely white, whereas the reverse displayed pale orange-yellow coloration with brown pigmentation. Ascomata were superficial, subglobose to ovate, ostiolate, and covered with terminal hairs. Asci were fusiform, irregularly shaped, eight-spored; spore-bearing parts measured (32.8–)35.6(–40.0) × (12.0–)16.0(–19.1) μm (n = 6). Ascospores were olive-brown at maturity, ovate to coffee bean-shaped with a central depression, (9.5–)12.8(–14.1) × (5.2–)6.6(–8.5) μm (n = 20).

Specimen examined: Korea, Jeollabuk-do, Buan-gun, 35°38′12.44″ N 126°27′42.10″ E, 21 Mar. 2025, isolated from S. thunbergii, strain KNUE 25S025 (NIBRFGC000513986); GenBank: PX121697 (ITS), PX127656 (TUB2), PX127657 (RPB2).

Notes: P. perlucidum was first described by Sergeeva (1956) as Chaetomium perlucidum from dead herbaceous stems and was transferred to Parachaetomium by Wang & Houbraken in 2022 [39,40]. The protologue described superficial, smoke-grey, subglobose to ovate, ostiolate ascomata; fusiform asci with spore-bearing parts 28–38 × 12–14 μm; and olivaceous, fusiform to elongated ovoid ascospores (11–14.5 × 5.5–7.5 μm) with a subapical or oblique germ pore [40]. The morphological features of strain KNUE 25S025 were consistent with these descriptions.

Hypomontagnella spongiphila Kuhnert, Fungal diversity 106: 23 (2020) [MB#833748] (Figure 10)

Figure 10.

Figure 10.

Open in a new tab

Morphology of Hypomontagnella spongiphila KNUE 24S533. A-B Colonies after 7 days of growth at 25 °C. The left side shows the front view and the right side shows the reverse view, on malt extract agar (A), and potato dextrose agar (B). Mononematous conidiophore with monoblastic conidiogenous cells and conidia (C), Synnematous conidiophore with polyblastic conidiogenous cells bearing clusters of conidia (D), Conidia (E). Scale bars: C-E = 10 μm.

Morphological characteristics: After 7 days of incubation at 25 °C in the dark, colonies on MEA reached 33.0–36.0 mm in diameter. Colonies were circular, cottony, white on the surface; the reverse was similar to the obverse. On PDA, colonies resembled those on MEA, with diameters of 40.0–44.0 mm. Conidiophores were mononematous or synnematous, hyaline to pale brown, and smooth- to rough-walled. Conidiogenous cells were predominantly polyblastic, although monoblastic development was occasionally observed. Conidia were globose to ovoid or clavate, (9.5–)12.8(–14.1) × (5.2–)6.6(–8.5) μm (n = 20).

Specimen examined: Korea, Jeju-do, Seongsan-eup, 33°27′36.868″ N 126°56′5.136″ E, 1 Aug. 2024, isolated from S. thunbergii, strain KNUE 24S533 (NIBRFGC000513997); GenBank: PX129499 (ITS), PX129500 (LSU), PX131339 (RPB2), PX131338 (TUB2).

Notes: H. spongiphila CLL 205 was initially identified as H. monticulosa together with the holotype strain MUCL 54604 in 2018, due to high sequence similarity and comparable metabolite production [41,42]. However, subsequent genomic comparisons demonstrated that strain CLL 205 represented a distinct species. Unlike MUCL 54604, which was isolated as a plant endophyte, CLL 205 was recovered from the marine sponge Sphaerocladina sp., from which the epithet “spongiphila” was derived [42]. Conidia of strain KNUE 24S533 were comparatively more rounded and smaller in size than those of H. monticulosa [41].

Palmiascoma acericola L.W. Li & Jian K. Liu, Mycosphere 14 (1): 1450 (2023) [MB#849144] (Figure 11)

Figure 11.

Figure 11.

Open in a new tab

Morphology of Palmiascoma acericola KNUE 25S034. A-B Colonies after 7 days of growth at 25 °C. The left side shows the front view and the right side shows the reverse view, on malt extract agar (A), and potato dextrose agar (B). Conidiomata observed on the colony surface on PDA (C). Section through conidiomata (D). Conidiogenous (E). Mature and immature conidia (F). Scale bars: C = 500 μm, D = 50 μm E = 10 μm F = 5 μm.

Morphological characteristics: After 7 days of incubation at 25 °C in the dark, colonies on MEA reached 47.0–50.0 mm in diameter. Colonies were dense and circular, with a flat elevation and a slightly rough surface; the colony surface was predominantly white. The reverse displayed a similar appearance to the surface, except for the central area, which exhibited a yellowish-gray coloration. On PDA medium, colonies reached 37.0–40.0 mm in diameter. Colonies were dense and circular, with a flat to slightly raised profile and a somewhat rough surface; the surface was entirely white, while the reverse was whitish-yellow with a gray center that became progressively darker toward the middle. Conidiomata were pycnidial, immersed in agar or superficial, appearing as black dots and containing dark pigmentation. Conidia were solitary, one-celled, oblong to ellipsoidal with rounded or obtuse ends, initially hyaline, becoming brown at maturity, aseptate, and smooth-walled, (3.0–)4.6(–6.2) × (2.1–)2.7(–3.1) μm (n = 20).

Specimen examined: Korea, Jeollabuk-do, Buan-gun, 35°37′6.08″ N 126°27′58.36″ E, 21 Mar. 2025, isolated from S. thunbergii, strain KNUE 25S034 (NIBRFGC000514079); GenBank: PX123871 (ITS), PX121739 (LSU), PX121773 (SSU), PX127658 (RPB2).

Notes: Palmiascoma acericola was originally collected as a saprobe from decaying branches of Acer truncatum, from which its epithet “acericola” is derived. The holotype was reported to represent only the sexual morph [43]. In contrast, strain KNUE 25S034 produced an asexual morph in culture, forming pycnidial conidiomata and solitary aseptate conidia. Although this feature was not described in the protologue, multilocus phylogenetic analyses confirmed its identity as P. acericola.

Aspergillus clavatophorus Sklenar, S.W. Peterson & Hubka, stud. Mycol. 88: 187 (2017) [MB#818936] (Figure 12)

Figure 12.

Figure 12.

Open in a new tab

Morphology of Aspergillus clavatophorus KNUE 25S318. A-B Colonies after 14 days of growth at 25 °C. The left side shows the front view and the right side shows the reverse side, on 25ppt malt extract agar (A), and 25ppt potato dextrose agar (B). Conidiophore (C), Conidia (D). Scale bar: C = 20 μm, D = 10 μm.

Morphological characteristics: After 14 days of incubation at 25 °C in the dark, colonies on 25 ppt MEA (malt extract agar supplemented with 25 ppt seawater) reached 12.0–15.0 mm in diameter. Colonies were circular, with a slightly filiform margin, a wrinkled surface, and a cottony texture; the colony surface was light greenish yellow, and the reverse appeared similar. On 25 ppt PDA (potato dextrose agar supplemented with 25 ppt seawater), colonies reached 11.0–14.0 mm in diameter. Colonies were circular, with a wrinkled surface and a cottony texture; the surface was light greenish yellow, and the reverse was dark brown at the center, becoming light yellowish brown toward the margin. Conidial heads were radiate, and conidiophores were uniseriate. Vesicles were spatulate to clavate, 11.0–14.9 μm wide (n = 5). Phialides were flask-shaped, 7.8–10.9 μm long. Conidia were initially smooth and ellipsoidal, later becoming rough-walled and subglobose, ovate, or barrel-shaped, (4.3–)5.1(–6.9) × (3.0–)4.0(–5.3) μm (n = 20).

Specimen examined: Korea, Jeju-do, Seongsan-eup, 33°24′07.3″ N 126°54′20.4″ E, 11 Apr. 2025, isolated from S. thunbergii, strain KNUE 25S318 (NIBRFGC000514080); GenBank: PX129233 (ITS), PX131336 (TUB2), PX131337 (CaM), PX131335 (RPB2).

Notes: A. clavatophorus was originally described in 2017 as a xerophilic species of Aspergillus, characterized by rapid growth on media supplemented with 5–15% NaCl [44]. Similarly, strain KNUE 25S318 showed little to no growth on PDA and MEA without added salinity, but slow growth on media supplemented with 25 ppt seawater, consistent with its halotolerant profile.

4. Discussion

This study reports six previously unrecorded endophytic fungi—Botryotrichum murorum, Subramaniula cristata, Parachaetomium perlucidum, Hypomontagnella spongiphila, Palmiascoma acericola, and Aspergillus clavatophorus—isolated from the brown alga Sargassum thunbergii collected along the Korean coast. Species identification was confirmed through detailed morphological observations and multilocus phylogenetic analyses, revealing the hidden fungal diversity associated with S. thunbergii and offering new insights into the ecological roles of algal endophytes.

Three of the isolates identified in this study, B. murorum, S. cristata, and P. perlucidum, belong to the family Chaetomiaceae. Members of this family are typically reported from terrestrial habitats such as soil, litter, and decaying plant materials [37,40]. While some Chaetomiaceae have previously been reported as algal-associated endophytes, these species are documented here for the first time from a marine algal host [8,10,13,14,18]. Their occurrence in S. thunbergii thus expands the known species-level diversity of Chaetomiaceae in marine environments and highlights the capacity of this host to support multiple lineages within the family [45]. Chaetomiaceae fungi are also prolific producers of structurally diverse secondary metabolites, such as chaetoglobosins, chaetoquadrins, and orsellides, which are known for antimicrobial and cytotoxic activities [18,21,37]. Their presence in S. thunbergii therefore illustrates both ecological adaptability and potential as sources of bioactive metabolites with pharmaceutical and agricultural relevance in marine environments.

Hypomontagnella spongiphila was originally described from a marine sponge and is now recognized as a distinct member of the genus Hypomontagnella [42]. Members of this genus, along with other taxa in the Hypoxylaceae, are notable for producing unusual secondary metabolites, including sporothriolide derivatives and other cytotoxic compounds, which have potential chemotaxonomic relevance within the family [41,42,46]. The isolation of H. spongiphila from Sargassum thunbergii extends its host range from sponge to algal substrates, underscoring ecological plasticity within the genus and reinforcing its potential as a reservoir of structurally novel metabolites with pharmaceutical potential. Palmiascoma acericola was first reported as a saprobe from woody tissues of a maple tree [43]. In this study, it was identified as an endophyte of S. thunbergii, representing the first record of this genus from a marine algal host. This finding broadens the ecological range of Palmiascoma and demonstrates its ability to inhabit novel host environments beyond woody substrates. The holotype of Aspergillus clavatophorus was originally isolated in 1987 from moldy paper, and subsequent reports documented its occurrence on cardboard, paintings, and fermented products such as puerh tea [44]. Our study provides the first evidence of this species as an algal endophyte, thereby extending its known distribution from terrestrial and anthropogenic substrates to a marine host. This discovery emphasizes the adaptive versatility of the species and suggests the possibility of unrecognized metabolic capacities relevant to saline environments [13,14,16].

The identification of six distinct fungal taxa from S. thunbergii demonstrates that this alga harbors a complex and taxonomically diverse endophytic community [10,11]. These fungi are likely to contribute to the algal holobiont by producing secondary metabolites that enhance host resilience against microbial pathogens, grazers, and abiotic stressors such as salinity and temperature fluctuations [21]. At the same time, the taxonomic novelty and ecological breadth of the isolates underscore their potential as valuable sources of bioactive compounds with practical applications [6,9,13,14,16,18].

Research on seaweed-associated fungi in Korea remains at an early stage, but the present study contributes to a growing body of evidence that coastal macroalgae represent a promising yet underexplored reservoir of fungal diversity [10,11,22]. Future work integrating ecological surveys, metabolite profiling, and genomic approaches will be critical for elucidating the functional roles of these fungi and for assessing their applied potential. Continued exploration of S. thunbergii and related brown algae along Korea’s coasts will likely uncover additional novel taxa that not only enrich fungal taxonomy but also hold promise for pharmaceutical, agricultural, and environmental applications.

Funding Statement

This work was supported by a grant from the National Institute of Biological Resources (NIBR202502103), funded by the ministry of Environment (MOE) of the Republic of Korea.

Disclosure statement

No potential conflict of interest was reported by the author(s).

References

  • 1.Kim S, Choi SK, Van S, et al. Geographic differentiation of morphological characteristics in the brown seaweed Sargassum thunbergii along the Korean coast: a response to local environmental conditions. JMSE. 2022;10(4):549. doi: 10.3390/jmse10040549. [DOI] [Google Scholar]
  • 2.Liu F-L, Li J-J, Liang Z-R, et al. A concise review of the brown seaweed Sargassum thunbergii—a knowledge base to inform large-scale cultivation efforts. J Appl Phycol. 2021;33(6):3469–3482. doi: 10.1007/s10811-021-02557-2. [DOI] [Google Scholar]
  • 3.Kang JY, Khan MNA, Park NH, et al. Antipyretic, analgesic, and anti-inflammatory activities of the seaweed Sargassum fulvellum and Sargassum thunbergii in mice. J Ethnopharmacol. 2008;116(1):187–190. doi: 10.1016/j.jep.2007.10.032. [DOI] [PubMed] [Google Scholar]
  • 4.Wu H, Liu H, Zhan D, et al. Research present situation of Sargassum thunbergii. Terr Nat Resour Stud. 2010;1:95–96. [Google Scholar]
  • 5.Wang X, Zhan D, Li M, et al. Preliminary studies on the nitrogen and phosphorus absorption capability of macroalgae. Prog Fish Sci. 2011;32:67–71. [Google Scholar]
  • 6.Ji N-Y, Wang B-G.. Mycochemistry of marine algicolous fungi. Fungal Divers. 2016;80(1):301–342. doi: 10.1007/s13225-016-0358-9. [DOI] [Google Scholar]
  • 7.Wilson D. Endophyte: the evolution of a term, and clarification of its use and definition. Oikos. 1995;73(2):274–276. doi: 10.2307/3545919. [DOI] [Google Scholar]
  • 8.Ahamed F, Murugan M.. Isolation and characterization of marine endophytic fungi from seaweeds, and bioactivity of their crude extracts. J Pure Appl Microbiol. 2019;13(3):1451–1460. doi: 10.22207/JPAM.13.3.15. [DOI] [Google Scholar]
  • 9.Teixeira TR, dos Santos GS, Armstrong L, et al. Antitumor potential of seaweed derived-endophytic fungi. Antibiotics. 2019;8(4):205. doi: 10.3390/antibiotics8040205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Kim J-W, Kim Y-J, Eom A-H.. Diversity and community structure of endophytic fungi isolated from the brown seaweed Sargassum thunbergii in Coastal Regions of Korea. Mycobiology. 2024;52(5):317–323. doi: 10.1080/12298093.2024.2416730. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Kim J-W, Kim Y-J, Eom A-H.. Morphological and phylogenetic characterization of endophytic fungi isolated from brown algae (Phaeophyceae) in Korea. Mycobiology. 2025;53(5):620–628. doi: 10.1080/12298093.2025.2535776. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Eo J-K, Choi M-S, Eom A-H.. Diversity of endophytic fungi isolated from Korean ginseng leaves. Mycobiology. 2014;42(2):147–151. doi: 10.5941/MYCO.2014.42.2.147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Suryanarayanan TS. Fungal endosymbionts of seaweeds. In: Jones EBG, Hyde KD, editors. Biology of marine fungi, progress in mycological research. Berlin, Heidelberg: Springer; 2011. p. 53–69. [DOI] [PubMed] [Google Scholar]
  • 14.Calado ML, Silva J, Alves C, et al. Marine endophytic fungi associated with Halopteris scoparia (Linnaeus) Sauvageau as producers of bioactive secondary metabolites with potential dermocosmetic application. PLoS One. 2021;16(5):e0250954. doi: 10.1371/journal.pone.0250954. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 15.Flewelling AJ, Johnson JA, Gray CA.. Antimicrobials from the marine algal endophyte Penicillium sp. Nat Prod Commun. 2013;8(3):373–374. 1934578X1300800324. [PubMed] [Google Scholar]
  • 16.Sahoo S, Subban K, Chelliah J.. Diversity of marine macro-algicolous endophytic fungi and cytotoxic potential of Biscogniauxia petrensis metabolites against cancer cell lines. Front Microbiol. 2021;12:650177. doi: 10.3389/fmicb.2021.650177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17.Sharma VK, Shah MP, Parmar S, et al. Fungi bio-prospects in sustainable agriculture, environment and nano-technology: volume 3: fungal metabolites, functional genomics and nano-technology. London, UK: Academic Press; 2021. [Google Scholar]
  • 18.Singh VK, Dwivedy AK, Singh A, et al. Fungal endophytes from seaweeds: an overview. In: Dubey AK, Singh VK, Singh RK, editors. Microbial biotechnology: volume 2. Application in food and pharmacology. Singapore: Springer; 2018. p. 483–498. [Google Scholar]
  • 19.Jones EBG. Fifty years of marine mycology. Fungal Divers. 2011;50(1):73–112. doi: 10.1007/s13225-011-0119-8. [DOI] [Google Scholar]
  • 20.Sarasan M, Puthumana J, Job N, et al. Marine algicolous endophytic fungi-a promising drug resource of the era. J Microbiol Biotechnol. 2017;27(6):1039–1052. doi: 10.4014/jmb.1701.01036. [DOI] [PubMed] [Google Scholar]
  • 21.Noor S, Begum M, Rony SR, et al. Bioactivity and chemical screening of endophytic fungi associated with seaweeds Gracilaria sp. and Sargassum sp. of the Bay of Bengal, Bangladesh. Sci Rep. 2025;15:1–17. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Kim J-W, Kim Y-J, Eom A-H.. Endophytic fungi isolated from the marine macroalga Dictyopteris pacifica in Korea. Mycobiology. 2025;53(1):47–56. doi: 10.1080/12298093.2024.2440975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Sarasan M, Job N, Puthumana J, et al. Exploration and profiling of hidden endophytic mycota of marine macroalgae with potential drug leads. FEMS Microbiol Lett. 2020;367:fnaa078. doi: 10.1093/femsle/fnaa078. [DOI] [PubMed] [Google Scholar]
  • 24.Erbert C, Lopes AA, Yokoya NS, et al. Antibacterial compound from the endophytic fungus Phomopsis longicolla isolated from the tropical red seaweed Bostrychia radicans. Botanica Marina. 2012;55(4):435–440. doi: 10.1515/bot-2011-0023. [DOI] [Google Scholar]
  • 25.Kjer J, Debbab A, Aly AH, et al. Methods for isolation of marine-derived endophytic fungi and their bioactive secondary products. Nat Protoc. 2010;5(3):479–490. doi: 10.1038/nprot.2009.233. [DOI] [PubMed] [Google Scholar]
  • 26.Cha J-E, Kim Y-J, Kim J-W, et al. Two new records of endophytic fungi isolated from Lindera obtusiloba in Korea: Colletotrichum citricola and Valsa ceratophora. Kor J Mycol. 2024;52:135–143. [Google Scholar]
  • 27.Gardes M, Bruns T.. ITS primers with enhanced specificity for basidiomycetes‐application to the identification of mycorrhizae and rusts. Mol Ecol. 1993;2:113–118. [DOI] [PubMed] [Google Scholar]
  • 28.White TJ, Bruns T, Lee S, et al. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis M, Gelfand D, editors. PCR protocols: a guide to methods and applications. London: Academic Press; 1990. p. 315–322. [Google Scholar]
  • 29.O’donnell K. Fusarium and its near relatives. In: Reynolds DR, Taylor JW, editors. The fungal holomorph: mitotic, meiotic and pleomorphic speciation in fungal systematics. Wallingford, UK: CAB International; 1993. [Google Scholar]
  • 30.Glass NL, Donaldson GC.. Development of primer sets designed for use with the PCR to amplify conserved genes from filamentous ascomycetes. Appl Environ Microbiol. 1995;61(4):1323–1330. doi: 10.1128/aem.61.4.1323-1330.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.O’Donnell K, Cigelnik E.. Two divergent intragenomic rDNA ITS2 types within a monophyletic lineage of the fungus Fusarium are non orthologous. Mol Phylogenet Evol. 1997;7(1):103–116. doi: 10.1006/mpev.1996.0376. [DOI] [PubMed] [Google Scholar]
  • 32.Woudenberg J, Aveskamp M, De Gruyter J, et al. Multiple Didymella teleomorphs are linked to the Phoma clematidina morphotype. Persoonia. 2009;22(1):56–62. doi: 10.3767/003158509X427808. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33.Liu YJ, Whelen S, Hall BD.. Phylogenetic relationships among ascomycetes: evidence from an RNA polymerse II subunit. Mol Biol Evol. 1999;16(12):1799–1808. doi: 10.1093/oxfordjournals.molbev.a026092. [DOI] [PubMed] [Google Scholar]
  • 34.Vilgalys R, Hester M.. Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species. J Bacteriol. 1990;172(8):4238–4246. doi: 10.1128/jb.172.8.4238-4246.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Hong S-B, Cho H-S, Shin H-D, et al. Novel Neosartorya species isolated from soil in Korea. Int J Syst Evol Microbiol. 2006;56(2):477–486. doi: 10.1099/ijs.0.63980-0. [DOI] [PubMed] [Google Scholar]
  • 36.Tamura K, Stecher G, Kumar S.. MEGA11: molecular evolutionary genetics analysis version 11. Mol Biol Evol. 2021;38(7):3022–3027. doi: 10.1093/molbev/msab120. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Wang X, Houbraken J, Groenewald JZ, et al. Diversity and taxonomy of Chaetomium and chaetomium-like fungi from indoor environments. Stud Mycol. 2016;84(1):145–224. doi: 10.1016/j.simyco.2016.11.005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38.Ames L. New cellulose destroying fungi isolated from military material and equipment. Mycologia. 1949;41(6):637–648. doi: 10.1080/00275514.1949.12017807. [DOI] [Google Scholar]
  • 39.Sergeeva MK. New species of the genus Chaetomium. Mycologia. 1956;48:671–683. [Google Scholar]
  • 40.Wang X, Han P, Bai F, et al. Taxonomy, phylogeny and identification of Chaetomiaceae with emphasis on thermophilic species. Stud Mycol. 2022;101(1):121–243. doi: 10.3114/sim.2022.101.03. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 41.Lambert C, Wendt L, Hladki AI, et al. Hypomontagnella (Hypoxylaceae): a new genus segregated from Hypoxylon by a polyphasic taxonomic approach. Mycol Prog. 2019;18(1–2):187–201. doi: 10.1007/s11557-018-1452-z. [DOI] [Google Scholar]
  • 42.Wibberg D, Stadler M, Lambert C, et al. High quality genome sequences of thirteen Hypoxylaceae (Ascomycota) strengthen the phylogenetic family backbone and enable the discovery of new taxa. Fungal Divers. 2021;106(1):7–28. doi: 10.1007/s13225-020-00447-5. [DOI] [Google Scholar]
  • 43.Li W, Liang R, Dissanayake A, et al. Mycosphere Notes 413–448: dothideomycetes associated with woody oil plants in China. Mycosphere. 2023;14(1):1436–1529. doi: 10.5943/mycosphere/14/1/16. [DOI] [Google Scholar]
  • 44.Sklenář F, Jurjević Ž, Zalar P, et al. Phylogeny of xerophilic aspergilli (subgenus Aspergillus) and taxonomic revision of section Restricti. Stud Mycol. 2017;88:161–236. doi: 10.1016/j.simyco.2017.09.002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 45.Redmond S, Kim JK, Yarish C, et al. Culture of Sargassum in Korea: techniques and Potential for Culture in the US. Orono, ME Maine Sea Grant College Program; 2014. [Google Scholar]
  • 46.Krohn K, Ludewig K, Aust H-J, et al. Biologically active metabolites from fungi. 3. sporothriolide, discosiolide, and 4-epi-ethisolide-new furofurandiones from Sporothrix sp., Discosia sp., and Pezicula livida. J Antibiot (Tokyo). 1994;47(1):113–118. doi: 10.7164/antibiotics.47.113. [DOI] [PubMed] [Google Scholar]

Articles from Mycobiology are provided here courtesy of Korean Society of Mycology

RESOURCES