Korean Journal of Mycology (Kor. J. Mycol.)
Hae-Dam Kim1, Che-Yeon Park1, Sashini Subodha1, Seo-Ryeong Lee1, Seong-Keun Lim1, Seung-Yeol Lee1,2,*, and Hee-Young Jung1,2
1Department of Plant Medicine, Kyungpook National University, Daegu 41566, Korea
2Institute of Plant Medicine, Kyungpook National University, Daegu 41566, Korea
*Correspondence to leesy1123@knu.ac.kr
Korean Journal of Mycology (Kor J Mycol) 2026 June, Volume 54, Issue 2, pages 155-163.
https://doi.org/10.4489/kjm.2026.54.2.5
Received on April 16, 2026, Revised on June 02, 2026, Accepted on June 04, 2026, Published on June 30, 2026.
Copyright © The Korean Society of Mycology.
This is an Open Access article which is freely available under the Creative Commons Attribution-Non-Commercial 4.0 International License (CC BY-NC) (https://creativecommons.org/licenses/by-nc/4.0/).
Humicola phialophoroides, Humicola-like aleuriospores, Morphological characterization, Phialophora-like conidiophores, Phylogeny
Fungi, which are eukaryotic organisms, play a significant role in maintaining the balance of ecosystems and provide various ecological services, from nutrient cycling to symbiotic relationships with plants and animals [1]. The family Chaetomiaceae, which belongs to the phylum Ascomycota, substantially influences agriculture, ecosystems, biotechnology, and the health of living beings by producing a wide range of structurally diverse metabolites [2]. The genus Humicola, found in the family Chaetomiaceae, was first introduced by Traaen in 1914 for two species of H. fuscoatra and H. grisea [3]. Members of the genus Humicola possess diverse bioactive metabolites and produce thermostable enzymes, thereby exhibiting significant biotechnological and industrial potential [4–8]. More than 50 species of this genus have been described and most of these species are commonly found in soil, decomposing plant matter, caves, and indoor spaces [7,9]. Moreover, some species of this genus are mycoparasites that colonize and invade the hyphae or survival structures, i.e., sclerotia, of other fungal species [4,6,7]. Although some species can control plant diseases caused by other fungal species, such as Phytophthora capsici, a few species can cause infections in humans [10–12]. Certain species belonging to this genus also act as biosorbents for heavy metals [13–15]. In Korea, studies on the genus Humicola remain limited, and only a few species, including H. olivacea and H. alopallonella, have previously been reported [16–18]. Members of this genus are known to inhabit diverse ecological niches, including soil, decaying plant materials, and plant-associated environments.
The discovery of previously unreported Humicola species not only expands our understanding of fungal diversity but also highlights the importance of continued exploration in varied ecosystems in South Korea. By defining an unreported species, our study contributes to the broader understanding of the taxonomy, ecology, and potential biotechnological applications of fungi. This study reports the cultural and morphological characteristics of a previously unreported species, providing detailed information regarding its importance within the genus.
The fungal strain KNUF-25-YP1 was isolated from internal tissues of the root collar of wilted red pepper plants collected from a red pepper field located in Yeongyang-gun, Gyeongbuk Province. Collected tissues were directly cultured on potato dextrose agar (PDA; Difco, Detroit, MI, USA) and incubated at 25°C for 7 days. The fungal strain KNUF-25-YP1 was selected for further molecular analyses, as well as cultural and morphological evaluations. The isolate was preserved at the Korean Agricultural Culture Collection under the accession number KACC 411310.
The isolate was cultured on PDA (Difco, Detroit, MI, USA), oatmeal agar (OA; Difco, Detroit, MI, USA), malt extract agar (MEA; Difco, Detroit, MI, USA), corn meal agar (CMA; Difco, Detroit, MI, USA), and potato carrot agar (PCA; HiMedia, Mumbai, India) for morphological and cultural characterization [9,10]. The cultures were maintained for 7 days at 25°C in the dark, and various morphological characteristics, such as size, color, mycelial shape, and details of the colony, including morphological features such as conidiophores, aleuriospores, chlamydospores, phialides, and conidia were observed. Morphological characteristics were examined under a light microscope (BX-50; Olympus, Tokyo, Japan).
For molecular identification, total genomic DNA was extracted from the KNUF-25-YP1 strain using the HiGene™ Genomic DNA Prep Kit (Biofact, Daejeon, Korea), according to the manufacturer’s instructions. Phylogenetic analysis was conducted based on two non-coding regions, namely the internal transcribed spacer (ITS) regions and partial sequences of the large subunit 28S rRNA (LSU) gene, which were amplified using the primer pairs ITS1F/ITS4 and LR0R/LR5, respectively [19–21]. Successful amplification was confirmed through electrophoresis using 1.0% HP agarose gels (Biopure, Cambridge, MA, USA). The amplified products were purified utilizing ExoSAP-IT (Thermo Fisher Scientific, Waltham, MA, USA) and then submitted to Macrogen (Seoul, Korea) for sequencing.
The acquired sequences were analyzed for similarity using the Basic Local Alignment Search Tool (BLAST) available in the National Center for Biotechnology Information (NCBI) database (Table 1). Phylogenetic trees were constructed based on the combined sequences of the ITS regions and LSU gene, using the maximum likelihood (ML) method in MEGA X [22]. Evolutionary distances for the ML analysis were calculated using the Tamura-Nei model, with bootstrap values derived from 1,000 replications [22].
Table 1. List of species and GenBank accession numbers of their sequences used for the phylogenetic analyses in this study
| Species name | Strain | GenBank Accession Number | |
|---|---|---|---|
| ITS | LSU | ||
| Humicola phialophoroides | KNUF-25-YP1 | PZ256383 | PZ256400 |
| Humicola phialophoroides | CBS 125784T | NR171742 | MH875195 |
| Humicola fuscoatra | MTCC 6329 | EF550969 | EU257376 |
| Humicola floriformis | CBS 815.97T | NR172533 | MH862678 |
| Humicola floriformis | PSSCE 126 | PP851769 | PP851769 |
| Humicola ampulliella | IHEM 24695 | OW987292 | OW987292 |
| Humicola ampulliella | CBS 116735T | NR172956 | LT993568 |
| Humicola leptodermospora | CBS 120095T | NR172960 | LT993584 |
| Humicola cuyabenoensis | CBS 398.97T | NR160060 | LT993573 |
| Humicola degenerans | CBS 232.65T | NR172958 | LT993574 |
| Humicola homopilata | CBS 157.55T | NR168166 | KM655364 |
| Phialocephala lagerbergii | CBS 266.33 | NR119426 | MH866885 |
ITS: internal transcribed spacer regions; LSU: partial sequences of the large subunit 28S rRNA.
TType strain.
The strain used in this study is indicated in bold.
Five different media were used to culture fungal colonies at 25°C for 7 days. Colonies on PDA measured 45–47 mm in diameter, and the observed side was an effused circular colony with an entire margin, a cottony texture, and abundant white-colored aerial hyphae. The reverse side of the colony was initially light yellow and gradually became light brown with irradiation (Fig. 1A). Colonies on OA measured 58–60 mm in diameter and exhibited a circular form with a slightly filamentous, even margin. White-colored aerial hyphae with a cottony texture were observed on the upper side, and an opaque appearance was observed on the reverse side (Fig. 1B). The colonies on CMA measured 49–51 mm in diameter and appeared circular and flat with an entire margin. The colonies had translucent opacity with surface hyphae and exhibited white-colored mycelia throughout the media and yellowish white-colored mycelia at the radius. Brownish yellow-colored mycelia appeared around the central inoculation point on the reverse side (Fig. 1C). On MEA, the colonies measured 34–36 mm in diameter and had a slightly filamentous, flat, irregular margin. White-colored surface hyphae were observed on the upper side, while the reverse side of the colony was pale yellow (Fig. 1D). On PCA, the colonies measured approximately 55–57 mm in diameter. The colonies were circular, had a smooth margin, and were slightly filamentous at the edge. A cottony texture with abundant white aerial hyphae was noted on the front side. The color on the reverse side was initially pale yellow and later became deep yellow to light orange, with even coloration extending to the radius of the colony (Fig. 1E). Dichotomously branched conidiophores with a pigmented basal portion measured 22–87 μm in length (n = 30) from the basal cell to the top of the phialides (Fig. 1F). Phialides were terminal or lateral and measured 8.4–13.9 μm in length; they were 2.4–3.5 μm broad at the base and 0.8–1.4 μm broad at the apical collarette (n = 30; Fig. 1K and L). Conidia were hyaline, unicellular, and spherical; they measured 1.6–2.6 μm in diameter (n = 50; Fig. 1H). Some conidia were arranged along with hyphae and accumulated as a ball-like mass at the tip of the phialides (Fig. 1M). Aleuriospores were spherical, unicellular, 6.7–10.2 μm in diameter (n = 40), and pigmented with light brown to dark brown color (Fig. 1G). Intercalary chlamydospores were oblong or obovate, measuring 9.2–11.8 × 4.8–5.5 µm (length × width, L × W) (Fig. 1I and J).
Note: Comparing the strain KNUF-25-YP1 and Humicola phialophoroides CBS 125784T, both strains exhibit similar cultural characteristics on PDA media. Both strains produced colonies with entire margins and a floccose to cottony texture. In addition, the mycelial growth of KNUF-25-YP1 was larger than that of H. fuscoatra on OA, CMA, MEA, and PCA media. For the morphological characteristics, KNUF-25-YP1 exhibited hyaline, spherical conidia, dichotomously branched conidiophores with pigmented basal portions, phialides with apical collarettes, and intercalary chlamydospores, which were comparable to those of the type strain (Table 2).
Fig. 1. Cultural and morphological characteristics of Humicola phialophoroides KNUF-25-YP1. A–E: obverse and reverse sides of the colony at 25°C after 7 days on PDA, OA, CMA, MEA, and PCA, respectively; F: Dichotomously branched conidiophore; G: Aleuriospores; H: Conidia; I,J: Chlamydospores; K,L: Phialides; M: Mass of conidia arranged on the tip of the phialide. Scale bars = 10 μm.
Table 2. Comparison of cultural and morphological characteristics of Humicola phialophoroides KNUF-25-YP1 with those of reference strains
| Characteristic | Humicola phialophoroides KNUF-25-YP1a | Humicola phialophoroides CBS 125784Tb | Humicola fuscoatra CGMCC 3.13428c |
|---|---|---|---|
| Colony on PDA | 45–47 mm at 25°C for 7 days; Entire margin, floccose to cottony; reverse light yellow to light brown | 7 mm/day at 32°C; White with abundant aerial hyphae | N/A |
| Colony on OA | 58–60 mm at 25°C for 7 days; Even margin, cottony, floccose; reverse opaque | N/A | 29–35 mm; Entire, olivaceous; reverse pale olivaceous gray |
| Colony on CMA | 49–51 mm at 25°C for 7 days; Flat, entire margin, white to yellowish; reverse brownish yellow | N/A | 24–30 mm; Slightly crenate; reverse pale olivaceous gray with soluble pigment |
| Colony on MEA | 34–36 mm at 25°C for 7 days; Flat, irregular to slightly filamentous margin; reverse pale yellow | N/A | 18–24 mm; Entire to slightly lobate, grayish sepia to fuscous black; reverse grayish sepia |
| Colony on PCA | 55–57 mm at 25°C for 7 days; Margin smooth to slightly filamentous, floccose; reverse pale yellow | N/A | 17–23 mm; Entire, grayish sepia to olivaceous at the center; reverse similar |
| Conidia | 1.6–2.6 µm; Hyaline, spherical | 1.8–2.6 µm; Hyaline, spherical, clustered mass at the phialide apex | N/A |
| Conidiophore | 22–87 µm; Dichotomously branched, pigmented at the base | 31–72 µm; Dichotomously branched, pigmented at the base | N/A |
| Aleuriospore | 6.7–10.2 µm; Light brown to dark brown, spherical, unicellular | 8.8–12.5 µm; Dark colored, spherical, unicellular | 6.5–9 × 7–9 µm; Olivaceous brown, globose, subglobose to oblate |
| Phialide | 8.4–13.9 µm; Terminal or lateral, apical collarette present | 9.5–15.5 µm; Lageniform, attenuate sursum | N/A |
| Chlamydospore | 9.2–11.8 × 4.8–5.5 µm (L × W); Intercalary, oblong to obovate | 9.0–12.1 × 5.6 µm (L × W); Intercalary, oblong to obovate, dark colored | N/A |
PDA: potato dextrose agar; OA: oatmeal agar; CMA: corn meal agar; MEA: malt extract agar; PCA: potato carrot agar; L: length; W: width.
aFungal strain used in this study; bSource of description [10]; cSource of description [9]; TType strain; N/A: Not available.
For the molecular identification of the isolated fungal strain, total genomic DNA was amplified to obtain sequences for the ITS regions and LSU gene, yielding lengths of 558 and 795 bp, respectively. The ITS regions exhibited 99.81% similarity to H. phialophoroides CBS 125784T, whereas the LSU exhibited 99.75% similarity to H. phialophoroides. H. fuscoatra and H. homopilata exhibited 94.66% and 93.91% similarities, respectively, to the combined sequence of the ITS regions and LSU gene, indicating their close relationship to H. phialophoroides. Thus, they were closely related to strain KNUF-25-YP1. A phylogenetic tree was constructed using the ML method based on the concatenated sequence of the ITS regions and LSU gene (Fig. 2).
Fig. 2. Maximum likelihood tree based on combined dataset of partial sequences, internal transcribed spacer (ITS) regions, and partial sequences of the large subunit 28S rRNA (LSU) gene, illustrating the phylogenetic placement of the strain KNUF-25-YP1 within the genus Humicola. Bootstrap values greater than 70% (based on 1,000 replications) are shown at branching points. The strain isolated in this study is indicated in bold and red. The tree was rooted using Phialocephala lagerbergii CBS 266.33 as an outgroup. Scale bar = 0.02 substitutions per nucleotide position.
This study provides the first morphological and phylogenetic evidence of H. phialophoroides in Korea. It was first described as a hyphomycete fungus that could produce pigmented, thick-walled, single-celled spores that appeared terminally or laterally on hyphae or on minimally differentiated conidiophores [9]. A comprehensive study between some Chaetomium species and Humicola species was conducted based on phylogenetic analyses by Wang et al. [9]. Humicola, previously known as an asexual genus in the family Chaetomiaceae, has been recently redefined to be capable of producing not only asexual spores, conidia, and aleuriospores but also sexual spores [9]. Humicola species exhibit biotechnological and industrial potential and are a thriving source of distinct and structurally diversified metabolites with a wide range of bioactivities [4]. Cerebrosides, terpenoids, xanthones, xanthoquinones, depsides, and benzenediol lactones are the compounds that have been detected in Humicola species. These compounds exhibit significant bioactivities, including antioxidant, antimalarial, anticancer, antimicrobial, antidiabetic, antiallergic, anticoagulant, insecticidal, anticoccidial, antihuman immunodeficiency virus (anti-HIV), and diacylglycerol acyltransferase inhibitory activities [4,5]. Moreover, Humicola species can produce thermostable enzymes, such as cellulases, lipases, xylanases, feruloyl esterases, pectate lyases, and beta-xylosidases, which have biotechnological and industrial importance [4,8]. Some Humicola species can also cause allergic reactions. For instance, H. pulvericola and H. sardiniae have been reported to cause fungal keratitis in the human eye [11,12]. In one study, H. fuscoatra, isolated from a sediment sample, was found to produce secondary metabolites that reactivated latent HIV type-1 (HIV-1) expression in an in vitro model [23]. Some species within the genus Humicola produce silver nanoparticles on reacting with aqueous silver nitrate solutions, resulting in the generation of extracellular nanoparticles with good dispersity and stability [24].
In the early stage of growth, H. phialophoroides forms branched Phialophora-like conidiophores bearing phialides with saucer-shaped tops. In the later stage, it produces Humicola-like aleuriospores [10]. Although several features of this species are similar to those of species belonging to the genera Phialophora and Humicola, their key fungal characteristics differ. H. phialophoroides is phylogenetically closer to the genus Humicola than to the genus Phialophora [10,25–28]. On preheating with sodium hydroxide, H. phialophoroides has been found to exhibit the greatest biosorption capacity for zinc and chromium [14,15]. So far, its pathogenicity in humans or plants has not been reported. Rather, it has been identified to have beneficial effects on the environment, including the potential suppression of fungal plant diseases, such as Phytophthora blight of pepper and leaf spot of spoon cabbage [10,13]. This fungus has a distinct ability to produce substances that can induce resistance against P. capsici by inhibiting zoospore germination or germ tube growth, with the active substances retained in the mycelium [13].
To the best of our knowledge, this is the first report of H. phialophoroides in Korea. In addition, although strain KNUF-25-YP1 was isolated from wilted red pepper plants, further studies are required to clarify its ecological role, including whether the species is saprophytic, endophytic, or associated with plant disease symptoms. The identification of this previously unreported species will contribute to the understanding and conservation of domestic biodiversity.
The authors declare no conflict of interest.
This work was carried out with the support of “Cooperative Research Program for Agriculture Science & Technology Development (Project No. RS-2025-02183115)” Rural Development Administration, Republic of Korea.
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