https://orcid.org/0000-0001-5353-2586
Abstract
Eurotiales is a relatively large order of Ascomycetes, well-known for their ability to produce secondary metabolites with potential beneficial applications. To understand their diversity and distribution, different environmental sources including soil, freshwater, insect, and indoor air were investigated. Eight strains of Eurotiales were isolated and identified based on their morphological characters and a multi-gene phylogenetic analysis of the ITS, BenA, CaM, and RPB2 regions. We identified eight taxa that were previously not reported from Korea: Aspergillus baeticus, A. griseoaurantiacus, A. spinulosporus, Penicillium anthracinoglaciei, P. labradorum, P. nalgiovense, Talaromyces atroroseus, and T. georgiensis. Detailed descriptions, illustrations, and phylogenetic tree for the eight new records species are presented, and information regarding the records is also discussed.
1. Introduction
Eurotiales is a relatively large order of Ascomycetes, which has both positive and negative impacts on human activities. The positive aspects include their utilization in food fermentation and biotechnology for the production of enzymes, organic acids, and medications [Citation1–4]. The negative aspects include opportunistic infections, indoor growth, food spoilage, and mycotoxin production [Citation1,Citation5,Citation6].
Eurotiales comprises 28 genera; 15 of them are classified under the Aspergillaceae family (Aspergillago, Aspergillus, Evansstolkia, Hamigera, Leiothecium, Monascus, Penicilliopsis, Penicillium, Phialomyces, Pseudohamigera, Pseudopenicillium, Sclerocleista, Warcupiella, Xerochrysium, and Xeromyces), two under Elaphomycetaceae (Elaphomyces and Pseudotulostoma), eight under Trichocomaceae (Acidotalaromyces, Ascospirella, Dendrosphaera, Rasamsonia, Sagenomella, Talaromyces, Thermomyces, Trichocoma), two under Thermoascaceae (Paecilomyces, Thermoascus), and one under Penicillaginaceae (Penicillago) [Citation1].
The genera Aspergillus, Penicillium, and Talaromyces (Eurotiomycetes, Eurotiales) are considered to be among the most chemically inventive fungi, producing a wide array of secondary metabolites [Citation1,Citation2,Citation7–9]. They are known for their ability to produce enzymes, organic acids, and antibacterial, anticancer, antifungal, antioxidative, and antiproliferative compounds [Citation1,Citation7,Citation8,Citation10–15]. In addition, they are effective biocontrol agents [Citation16].
The genus Aspergillus was first described by Micheli [Citation17]. Members of this genus are usually found in soil, water, decaying vegetation, seeds, grains, and indoor air environment [Citation18–20]. Currently, the genus Aspergillus is divided into six subgenera, 27 sections, 75 series, and 446 species [Citation1].
The genus Penicillium was erected by Link [Citation21]. Members of the genus are isolated from diverse substrates, including soil, water, air, indoor environments, and food products [Citation22–24]. Currently, this genus is divided into two subgenera, 32 sections, 89 series, and 483 accepted species [Citation1].
The genus Talaromyces was initially described by Benjamin as a sexual state of the genus Penicillium [Citation25]. Talaromyces species are distributed worldwide and are commonly isolated from soil, water, indoor environment, and human [Citation8,Citation26–29]. Currently, the genus Talaromyces is divided into eight sections and 171 accepted species [Citation1,Citation8,Citation27].
Although Aspergillus, Penicillium, and Talaromyces species are of economic importance, the current number of the three genera found in Korea is still limited [Citation23,Citation28–31]. These genera are relevant and significant in human life, thus it is important to isolate and identify them.
This work presents eight new records of Aspergillus baeticus, A. griseoaurantiacus, A. spinulosporus, Penicillium anthracinoglaciei, P. labradorum, P. nalgiovense, Talaromyces atroroseus, and T. georgiensis in Korea based on both morphological characteristics and phylogenetic analyses.
2. Materials and methods
2.1. Sample collection, and isolation
Soil samples were collected from Kunryang-ri, Cheongyang, Chungnam Province, Samgak-dong, Buk-gu, Gwangju, and Sangju, Gyeongsang Province in South Korea. Freshwater and insects samples were collected from Hanbat Arboretum located in Daejeon, and Kunryang-ri, Cheongyang, Chungnam Province. The samples were kept at 4 °C until further use. Fungal isolations from soil, freshwater, and insect were carried out as previously detailed [Citation28, Citation32]. Strains were isolated using potato dextrose agar (PDA; Becton, Dickinson and Co., Sparks, MD) and malt extract agar (MEA; Becton, Dickinson and Co., Sparks, MD) amended with 50 ppm of the antibiotic neomycin. For indoor air, colonies were picked from the contaminated plates kept at 25 °C. Individual colonies with various morphologies were picked and transferred to new PDA plates. All the pure isolates were maintained in PDA slant tubes and 20% glycerol at −80 °C at the Environmental Microbiology Laboratory Fungarium, Chonnam National University, Gwangju, South Korea. The strains were also deposited at the Culture Collection of the Nakdonggang National Institute of Biological Resources (NNIBR), Sangju, South Korea under the numbers NNIBRFG46707 and NNIBRFG46718, and the Collection of the National Institute of Biological Resources (NIBR), Incheon, Korea under the numbers NIBRFGC000508631, NIBRFG0000509598, NIBRFG0000509600, EKFTFGC000000138, and EKFTFGC000000142, and YYMAFGC000000144.
2.2. DNA extraction, PCR, and sequencing
Genomic DNA extractions were performed on seven-day-old colonies grown on PDA using the Solg™ Genomic DNA Prep Kit (Solgent Co. Ltd., Daejeon, South Korea). The internal transcribed spacer (ITS) rDNA, β-tubulin (BenA), calmodulin (CaM), and RNA polymerase II second largest subunit (RPB2) gene regions were amplified by PCR using the primer pairs ITS5/ITS4 [33] and V9G/ITS4 [Citation33,Citation34], Tub2Fd/Tub4Rd [Citation35] and T1/T22 [Citation36], CF1L/CF4 [Citation36], Cmd5/Cmd6 [Citation37], and RPB2-5F/RPB2-7cR [Citation38], respectively. PCR was done in 20 μL volumes with AccuPower PCR PreMix (BioneerCorp., Daejeon, South Korea), 2 μL genomic DNA, 1.5 μL of each primer (5 pmol/μL), and 14 μL deionized water. The PCR thermal cycle programs for ITS, BenA, CaM, and RPB2 amplification were as follows: initial denaturing step of 95 °C for 5 min, followed by 35 cycles of denaturation at 95 °C for 30 s, annealing at 52 °C (ITS) or 55 °C (BenA, CaM, RPB2) for 30 s, elongation at 72 °C for 45 s, and final extension at 72 °C for 7 min. The PCR products were then purified using an Accuprep PCR Purification Kit (BioneerCorp., Daejeon, South Korea). The purified PCR products were sequenced in both directions with the same primers by Macrogen (Daejeon, South Korea).
2.3. Molecular analyses
The taxa used in the phylogenetic analysis were obtained from previous studies [Citation1, Citation28, Citation39,Citation40], and downloaded from GenBank (www.ncbi.nlm.nih.gov/genbank/). SeqMan v. 7.0.0 (DNAstar, Madison, WI) was used to assemble the consensus sequences. The sequences were aligned using the MAFFT v.7 online program (http://mafft.cbrc.jp/alignment/server/) [Citation41] and manually optimized using MEGA v.7 [Citation42]. Maximum-likelihood (ML) analysis was performed using the RAxML-HPC2 on XSEDE (v.8.2.12) in the CIPRES Science Gateway (https://www.phylo.org/portal2) using a GTRGAMMA model with rapid bootstrap analysis followed by 1000 bootstrap replicates. The consensus trees were viewed in FigTree v. 1.3.1 [Citation43]. The newly generated sequences in this study were deposited in GenBank ().
Table 1. Fungal species and sequences used in phylogenetic analyses.
2.4. Morphological studies
Methods used for morphological observations followed previous study [Citation8, Citation44–46]. Colony characteristics were recorded after seven days of incubation. Micro-morphological characters were observed under a light microscope using a differential interference contrast microscope (Olympus BX53, Tokyo, Japan) and Olympus DP74 digital camera.
3. Results
3.1. Phylogenetic analyses
The phylogenetic relationship of isolates CNUFC SF23, CNUFC CY2264, and CNUFC GRS15 with accepted Aspergillus species was determined by analysis of concatenated sequence datasets of four loci (ITS, BenA, CaM, and RPB2). The concatenated alignment consisted of 2748 nucleotides, including inserted gaps (ITS: 675 bp, BenA: 459 bp, CaM: 603 bp, and RPB2: 1011 bp).
The phylogenetic relationship of isolates CNUFC CY2234, CNUFC MOP1, and CNUFC CY224 with accepted Penicillium species was determined by analysis of concatenated sequence datasets of four loci (ITS, BenA, CaM, and RPB2). The concatenated alignment consisted of 2619 nucleotides, including inserted gaps (ITS: 584 bp, BenA: 499 bp, CaM: 581 bp, and RPB2: 955 bp).
The phylogenetic relationship of isolates CNUFC SJ322 and CNUFC DW211 with accepted Talaromyces species was determined by analysis of concatenated sequence datasets of four loci (ITS, BenA, CaM, and RPB2). The concatenated alignment consisted of 2806 nucleotides, including inserted gaps (ITS: 653 bp, BenA: 573 bp, CaM: 694 bp, and RPB2: 886 bp).
Isolates CNUFC GRS15, CNUFC SF23, and CNUFC CY2264 were clustered with the ex-type strain of A. baeticus, A. griseoaurantiacus, and A. spinulosporus, respectively, with strong statistical support (). In , isolates CNUFC CY2234, CNUFC MOP1, and CNUFC CY224 were clustered with the ex-type strain of P. anthracinoglaciei, P. labradorum, and P. nalgiovense with 100% ML support, respectively. Isolates CNUFC SJ322 and CNUFC DW211 were grouped with the ex-type strain of T. atroroseus, and T. georgiensis, respectively, with strong statistical support ().
Figure 1. Maximum-likelihood (RAxML) analysis based on combined ITS, BenA, CaM, and RPB2 sequence data showing the relationship of the isolates CNUFC SF23, CNUFC CY2264, and CNUFC GRS15 with related species in sections Aenei, Nidulantes, and Usti of the genus Aspergillus. The numbers above or below the branches represent maximum-likelihood bootstrap percentages. Bootstrap values equal to or greater than 70% are indicated above or below the branches. Aspergillus ochraceoroseus NRRL 28622 was used as the outgroup. The newly generated sequence is indicated in blue.
Figure 2. Maximum-likelihood (RAxML) analysis based on combined ITS, BenA, CaM, and RPB2 sequence data showing the relationship of the isolates CNUFC CY2234, CNUFC MOP1, and CNUFC CY224 with related species in sections Brevicompacta, Canescentia, Chrysogena, Exilicaulis, and Robsamsonia of the genus Penicillium. The numbers above or below the branches represent maximum-likelihood bootstrap percentages. Bootstrap values equal to or greater than 70% are indicated above or below the branches. Aspergillus glaucus NRRL 116 was used as the outgroup. The newly generated sequence is indicated in blue.
Figure 3. Maximum-likelihood (RAxML) analysis based on combined ITS, BenA, CaM, and RPB2 sequence data showing the relationship of the isolates CNUFC SJ322, and CNUFC DW211 with species in sections Helici, Purpurei, Talaromyces, Tenues, and Trachyspermi of the genus Talaromyces. The numbers above or below the branches represent maximum-likelihood bootstrap percentages. Bootstrap values equal to or greater than 70% are indicated above or below the branches. Trichocoma paradoxa CBS 103.73 was used as the outgroup. The newly generated sequence is indicated in blue.
3.2. Taxonomy
Aspergillus baeticus A. Novakova & Hubka, Int. J. Syst. Evol. Microbiol. 62 (Pt.2): 2783 (2012) [MB#564188] ():
Figure 4. Morphology of Aspergillus baeticus CNUFC GRS15. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G–I) Conidiophores; (J) Conidia. Scale bars = 10 µm.
Description: On CYA at 25 °C, colonies slightly sulcate; mycelium white; texture floccose at center; sporulation sparse to moderate; no pigment or exudate produced; reverse yellow-brown at center to grayish yellow at margin, and reached 23–25 mm in diameter after seven days. On MEA at 25 °C, colonies raised at center, texture floccose; sporulation strong; no pigment or exudate produced; reverse moderate yellow, and reached 36–40 mm in diameter after seven days. On YES at 25 °C, colonies slightly raised at center; radially sulcate; texture velutinous to slightly floccose; sporulation sparse to moderate; reverse vivid yellow and reached 34–37 mm in diameter after seven days.
Micromorphology: Stipes smooth walled, 105.5–491(–601.5) × 5–7.5 μm. Vesicles broadly elliptical or elongated, (9.5–)11–18 × 13–20.5(–23.5) μm. Metulae 5–6.5 × (2.5–)3–4 μm. Phialides 6.5–8.5(–9) × 2–3.5 μm. Conidia globose to subglobose, finely roughened, (2.5–)3–4.5 μm.
Material examined: Republic of Korea, Samgak-dong, Buk-gu, Gwangju (35°11’58.1”N 126°53’58.1”E), from soil, October 12, 2019 (culture CNUFC GRS15).
Notes: Our strain did not produce Hülle cells on CYA and grew slower than that of A. baeticus (ex-type strain) on CYA at 25 °C (23–25 mm vs. 35–38 mm) [Citation47].
Aspergillus griseoaurantiacus Visagie, Hirooka & Samson, Studies in Mycology 78: 112 (2014) [MB#809197] ():
Figure 5. Morphology of Aspergillus griseoaurantiacus CNUFC SF23. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G–I) Conidiophores; (J) Conidia. Scale bars = 10 µm.
Description: On CYA at 25 °C, colonies radially sulcate; mycelium white; colony texture cottony at center; sporulation moderate; reverse moderate yellow and reached 26–28 mm in diameter after seven days. On MEA at 25 °C, colonies raised at center; radially sulcate; margins entire; texture velvety; sporulation moderate; reverse vivid yellow and reached 25–27 mm in diameter after seven days. On YES at 25 °C, colonies radially sulcate; mycelium white; colony texture floccose; sporulation strong at center and sparse at margins; reverse pale yellow and reached 32–38 mm in diameter after seven days.
Micromorphology: Stipes smooth walled, 241–449.5 × 4.5–6 μm. Vesicles spathulate or elongated, 8.5–17.5 μm wide. Metulae 3–5.5(–7) × 2.5–4(–5) μm. Phialides ampulliform, 3 per metula, 4–7.5(–8.5) × 2–3 μm. Conidia globose to subglobose, some ellipsoidal, finely roughened, 2.5–3.5 × 2–2.5 μm.
Material examined: Republic of Korea, 54–30 Mukdong-gil, Cheongyang-eup, Cheongyang-gun, Chungcheongnam-do (33°14’52.7”N 126°24’48.3”E), from soil, February 2021, H.B. Lee (culture CNUFC SF23).
Notes: The size of metulae of our strain was slightly shorter than that of the ex-type of A. griseoaurantiacus (3–5.5 [–7] × 2.5–4 [–5] μm vs. 4–10 × 3–5.5 µm) [Citation48]. In addition, the ex-type of A. griseoaurantiacus did not produce the globose and subglobose conidia observed in our strain [Citation48].
Aspergillus spinulosporus Hubka, S.W. Peterson, M. Kolařík, Plant Systematics and Evolution 302 (9): 1290 (2016) ():
Figure 6. Morphology of Aspergillus spinulosporus CNUFC CY2264. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G, H) Conidiophores; (I) Conidia; (J) Hülle cells. Scale bars: G–I = 10 µm, J = 20 µm.
Description: On CYA at 25 °C, colonies raised at center; mycelium white; sporulation moderate; reverse deep orange at center and moderate orange yellow at margins and reached 29–33 mm in diameter after seven days. On MEA at 25 °C, colonies plane; poor sporulation; Hülle cells abundant; reverse pale yellow and reached 43–48 mm in diameter after seven days. On YES at 25 °C, colonies slightly raised at center; radially sulcate; mycelium white; colony texture floccose; sporulation absent; reverse light to moderate orange yellow and reached 25–30 mm in diameter after seven days.
Micromorphology: Hülle cells globose, subglobose to ovoid, 15–21.5 μm wide. Conidiophores with smooth stipes, hyaline to light brown, 181–383 × 5.5–8.5 μm. Vesicles subclavate, 10.5–16 μm wide. Metulae hyaline, 7.5–10 × 3–4.5 μm. Phialides hyaline, flask-shaped, 6.5–10 × 3–4 μm. Conidia globose, some oval, smooth, 3–4 μm.
Material examined: Republic of Korea, Kunryang-ri, Cheongyang-eup, Cheongyang, Chungnam Province (36°26’16.2”N 126°46’04.6”E), from soil, February 25, 2022, H.B. Lee (culture CNUFC CY2264).
Notes: There were differences in the size of conidiophore and Hülle cell when compared with the ex-type strain of A. spinulosporus described in Chen et al. [Citation49].
Penicillium anthracinoglaciei L. Perini, Frisvad & Zalar, Microbial Ecology 86, 282–296 (2022) [MB#835602] ():
Figure 7. Morphology of Penicillium anthracinoglaciei CNUFC CY2234. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G, H) Conidiophores; (I) Conidia. Scale bars= 20 µm.
Description: On CYA at 25 °C, colonies radially sulcate; mycelium white; colony texture velvety; sporulation strong; reverse moderate yellow and reached 28–30 mm in diameter after seven days. On MEA at 25 °C, mycelium white and green; texture velvety; sporulation moderate; reverse pale yellow and reached 16–18 mm in diameter after seven days. On YES at 25 °C, colonies raised at center, radially sulcate; mycelium white sometimes green; colony texture floccose; sporulation absent at center and sparse at margins; reverse pale yellow and reached 20–24 mm in diameter after seven days.
Micromorphology: Conidiophores terverticillate. Stipes 85–215 × 3–4.5 μm. Metulae 6–11 × 3–4.5 μm. Phialides 5–13 × 2–4.5 μm. Conidia subglobose to ellipsoidal, 2.5–4 × 2–3.5 μm.
Material examined: Republic of Korea, Kunryang-ri, Cheongyang-eup, Cheongyang, Chungnam Province (36°26’16.2”N 126°46’04.6”E), from dead bee in rainwater, July 2022, H.B. Lee (culture CNUFC CY2234).
Notes: Compared with the ex-type strain of P. anthracinoglaciei, our strain grew faster on CYA (28–30 mm vs. 13–20 mm) [Citation50].
Penicillium labradorum Gibas, Wiederh, C. Sanders, Rothacker, E.R. Rogers & Fales, Medical Mycology, 58(8): 1061 (2020) ():
Figure 8. Morphology of Penicillium labradorum CNUFC MOP1. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G, H) Conidiophores; (I) Conidia. Scale bars = 10 µm.
Description: On CYA at 25 °C, colonies wrinkled; mycelia white; sporulation poor; soluble pigments absent, exudates vivid yellow droplets; reverse moderate to strong yellow and reached 19–22 mm in diameter after seven days. On MEA at 25 °C, colonies plane; mycelia white, sporulation absent; reverse pale yellow and reached 17–22 mm in diameter after seven days. On YES at 25 °C, colonies sulcate; texture floccose, slightly raised at center, sporulation absent; reverse light to moderate yellowish brown and reached 13–21 mm in diameter after seven days.
Micromorphology: Conidiophores monoverticillate. Stipes 5.5–17.5 μm. Phialides smooth, ampulliform, 2–4 per stipe, 5–6.5(–7) × 2–3 μm. Conidia globose to subglobose, finely roughened, 2–2.5 μm.
Material examined: Republic of Korea, Gwangju, Buk-gu, Chonnam National University (35°10’34.0”N 126°54’21.0”E), from an indoor air sample, April 9, 2020 (culture CNUFC MOP1).
Notes: Compared with the ex-type strain of P. labradorum, there was a slight difference. Our strain grew faster on CYA (19–22 mm vs. 10–11 mm) and MEA (17–22 mm vs. 16–17 mm) [Citation39].
Penicillium nalgiovense Laxa, Zentralbl. Bakteriol. 2. Abt. 86 (5–7): 160 (1932) [MB#114239] ():
Figure 9. Morphology of Penicillium nalgiovense CNUFC CY224. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G–I) Conidiophores; (J) Conidia. Scale bars = 10 µm.
Description: On CYA at 25 °C, colonies radially sulcate; margins entire; mycelium white and greenish grey; colony texture velvety to floccose; sporulation moderate to strong; reverse grayish yellow and reached 29–30 mm in diameter after seven days. On MEA at 25 °C, colonies texture floccose, mycelia white; sporulation poor or absent; soluble pigments absent; reverse moderate to strong yellow and reached 26–30 mm in diameter after seven days. On YES at 25 °C, colonies raised at center; radially and concentrically sulcate; margins irregular; mycelia white; sporulation moderate; reverse grayish yellow and reached 26–29 mm in diameter after seven days.
Micromorphology: Conidiophores biverticillate, terverticillate, and quaterverticillate. Metulae 6–15 × 2–3 µm. Phialides flask-shaped, 4.5–12 × 2–2.5 µm. Conidia globose to subglobose, smooth, 2.5–3.5 µm in diam.
Material examined: Republic of Korea, Kunryang-ri, Cheongyang-eup, Cheongyang, Chungnam Province (36°26’16.2”N 126°46’04.7”E), from soil, May 15, 2022, H.B. Lee (culture CNUFC CY224).
Notes: The morphological characters, such as biverticillate, terverticillate, and quaterverticillate conidiophores, flask-shaped phialides, and globose to subglobose conidia, of our strain were similar to the ex-type strain of P. nalgiovense [Citation51].
Talaromyces atroroseus N. Yilmaz, Frisvad, Houbraken & Samson, PLoS One 8(12): e84102,8 (2013) ():
Figure 10. Morphology of Talaromyces atroroseus CNUFC SJ322. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G, H) Conidiophores; (I) Conidia. Scale bars: G = 20 µm, H, I = 10 µm.
Description: On CYA at 25 °C, colonies texture floccose and velvety; mycelia white and olive green; sporulation strong; soluble pigments red; reverse dark red and reached 31–34 mm in diameter after seven days. On MEA at 25 °C, colonies texture floccose, mycelia white to grayish olive green; sporulation strong; soluble pigments absent; reverse pale greenish yellow and reached 32–38 mm in diameter after seven days. On YES at 25 °C, colonies slightly raised at center, texture velvety; mycelia white; sporulation poor; no soluble pigments; reverse vivid reddish orange and reached 29–37 mm in diameter after seven days.
Micromorphology: Conidiophores biverticillate. Stipes (83.5–)94–137.5(–154) × 2–3.5 μm. Metulae 8.5–12 × 2–3.5 μm. Phialides acerose, 8–11.5 × 2–3 μm. Conidia finely rough to rough, ellipsoidal, 2.5–3 × 2–2.5 μm.
Material examined: Republic of Korea, Namseong-dong, Sangju-si, Gyeongsangbuk-do (36.4109°N, 128.1591°E), from soil, July 31, 2019 (culture CNUFC SJ322).
Notes: The morphological characters of our strain were similar to the ex-type strain of T. atroroseus [Citation52].
Talaromyces georgiensis M. Guevara-Suarez, D.A. Sutton & N. Wiederhold, Mycoses 60 (10): 656 (2017) [MB#820460] ():
Figure 11. Morphology of Talaromyces georgiensis CNUFC DW211. (A, D) Colonies on Czapek yeast autolysate agar (CYA); (B, E) malt extract agar (MEA); (C, F) yeast extract sucrose agar (YES) (A–C: obverse view and D–F: reverse view); (G–I) Conidiophores; (J) Conidia. Scale bars = 10 µm.
Description: On CYA at 25 °C, colonies raised at center; texture velvety; mycelium white; sporulation sparse; reverse yellowish white and reached 22–23 mm in diameter after seven days. On MEA at 25 °C, colonies cottony; mycelium white becoming greenish grey; sporulation moderate; reverse grayish yellow, and reached 31–33 mm in diameter after seven days. On YES at 25 °C, colonies cottony; mycelium white; margins entire; sporulation absent; exudates and soluble pigments absent; reverse pale yellow, and reached 23–26 mm in diameter after seven days.
Micromorphology: Conidiophores monoverticillate. Stipes rough-walled, 12.5–35.5(–37.5) x 2–3 μm. Phialides acerose, 2–4 per stipe, 7–13.5 × 2–3 μm. Conidia smooth to finely rough-walled, globose to subglobose, 2–3(–3.5) μm.
Material examined: Republic of Korea, Mannyeon-dong, Seo-gu, Daejeon East Dongsan of Hanbat Arboretum (36°22’06.4”N 127°23’24.9”E), from freshwater, February 15, 2020 (culture CNUFC DW211).
Notes: Compared with the ex-type strain of T. georgiensis, our strain grew slower on CYA (22–23 mm vs. 29–31 mm) and MEA (23–26 mm vs. 28–30 mm) [Citation26].
4. Discussion
Aspergillus, Penicillium, and Talaromyces species have been frequently isolated from various environments such as soil, water, air, seed, and food [Citation1,Citation8,Citation18,Citation19,Citation22]. However, isolation of fungi from insects is still limited. In this study, P. anthracinoglaciei species was isolated from insect sample (dead bee). They are of great interest to pharmacologists due to their ability to produce a wide range of biological activities, including anti-inflammatory, antibacterial, antioxidant, and anti-cancer properties. More than 315 bioactive metabolites of the genus Aspergillus and 221 compounds of the genus Talaromyces have been documented [Citation53,Citation54].
Eight isolates from different niches were identified as A. baeticus, A. griseoaurantiacus, A. spinulosporus, P. anthracinoglaciei, P. labradorum, P. nalgiovense, T. atroroseus, and T. georgiensis belonging to sections Usti, Nidulantes, Brevicompacta, Exilicaulis, Chrysogena, Trachyspermi, and Helici, respectively ().
Aspergillus spinulosporus acts as a biocontrol agent against Xanthomonas oryzae and produces signals that upregulate superoxide dismutase to protect plants. It also exhibits chitinolytic and amylolytic activities [Citation55]. Additionally, it causes infections in the central nervous system and is associated with unique microbiomes found in tumors [Citation56,Citation57]. Aspergillus spinulosporus has been isolated from patients with aspergillosis, birds, clinical samples, brain tissue, sputum, BAL samples, undetermined sources, and asthmatic patients [Citation56–61] and from soil in the current study. Aspergillus griseoaurantiacus exhibits the highest chitinase and chitosanase activities when cultured under solid-state fermentation of potato shell [Citation62]. Chitinase shows antifungal activity against the pathogenic fungus, Fusarium solani [Citation62]. Chitosan oligosaccharides, which are the degraded products of chitosan obtained through chitosanase, exhibit good antibacterial and antioxidant activities [Citation62]. Aspergillus griseoaurantiacus was isolated from house dust collected from Thailand, Mexico, and Yela of Kosrae Island [Citation48], as well as from a soil sample in this study. Aspergillus baeticus has been isolated from cave sediment, cave air and bat cadaver in Spain [Citation47,Citation63], and from soil environment in this study.
Penicillium anthracinoglaciei acts as decomposers or parasites of glacier ice algae by utilizing and converting the pigment purpurogallin carboxylic acid-6-O-β-d-glucopyranoside into purpurogallin carboxylic acid [Citation50]. The species was isolated from the surface of the Greenland Ice Sheet [Citation50] and from dead bee in rainwater in this study. Penicillium labradorum isolated from a Labrador Retriever with disseminated fungal disease shows some clinical signs, including lethargy, lymphadenopathy, tachypnea, moderate pitting edema, and non-weight bearing lameness in the right hind limb [Citation39]. Additionally, it was isolated from the bone tissue of a 6-year-old female Beagle dog [Citation64] and was identified as an indoor contaminant in this study. Penicillium nalgiovense has various applications in medicinal chemistry and organic synthesis. It is capable of producing isocoumarins such as dichlorodiaportin, diaportinol, and diaportinic acid [Citation65]. In addition, it is involved in coprogen, amphotericin B, and penicillin production, protease production, fermentation of salchichon, and serves as a starter culture in dry fermentation in the food industry [Citation66–70]. Penicillium nalgiovense was isolated from the soil from an abandoned penguin’s nest, from Ellischauer cheese [Citation51,Citation68], and from the soil in this study.
Talaromyces georgiensis has been isolated from clinical samples [Citation26], from animal joint fluid [Citation71], and a 4-year-old male Schnauzer dog [Citation64]. Our new isolate, T. georgiensis CNUFC DW211, was isolated from a freshwater habitat in Korea. Talaromyces atroroseus has been reported in respiratory specimens of patients with pulmonary disorders, house dust (from Mexico, Thailand, and South Africa), coprophilous, red sweet bell pepper, a contaminated Petri dish, mouse dung, soil, and as a parasite in Aspergillus niger culture [Citation8,Citation52,Citation72]. Talaromyces atroroseus from this study was isolated from soil sample. Talaromyces atroroseus secretes large amounts of red pigments that have great potential for coloring foods [Citation52] and many other compounds such as glauconic acid, glaucanic acid, Monascus red pigments, purpactins AC, purpuride, purpurogenone, N-glutarylmonascorubramine, monascorubrin, monascorubramine, rubropunctatin and ZG-1494α [Citation8,Citation52]. Therefore, future studies are required for the isolation and identification of compounds from Talaromyces atroroseus in Korea.
In this study, eight new records of Eurotiales were identified from different niches in Korea. Our results suggest that there are numerous undescribed species awaiting discovery in Korea. Further exploration and sampling could result in the discovery of fungal groups that could be utilized for various taxonomic and ecological research.
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References
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