A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China

Yin, Changzhun; Zhang, Zhaoxue; Wang, Shi; Liu, Wenwen; Zhang, Xiuguo

doi:10.3390/jof10100700

Open AccessArticle

A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China

by

Changzhun Yin

¹,

Zhaoxue Zhang

²

,

Shi Wang

¹,

Wenwen Liu

¹ and

Xiuguo Zhang

^1,2,*

¹

College of Life Sciences, Shandong Normal University, Jinan 250300, China

²

Shandong Provincial Key Laboratory for Biology of Vegetable Diseases and Insect Pests, College of Plant Protection, Shandong Agricultural University, Taian 271000, China

^*

Author to whom correspondence should be addressed.

J. Fungi 2024, 10(10), 700; https://doi.org/10.3390/jof10100700

Submission received: 25 July 2024 / Revised: 3 October 2024 / Accepted: 5 October 2024 / Published: 7 October 2024

Download

Browse Figures

Review Reports Versions Notes

Abstract

:

In an extensive fungal investigation conducted in southern China, a large number of fungal strains were isolated by collecting and treating diseased and decayed leaves. Using internal transcribed spacer regions (ITSs) sequence data for a BLAST search to screen for suspected strains of Daldinia, followed by phylogenetic analysis using internal transcribed spacer regions, partial sequences of the large subunit of the rDNA (LSU), RNA polymerase II (rpb2), and beta tubulin (tub2) sequence data, combined with morphological characteristics of anamorphic species, ninety-four strains of Daldinia were identified. Furthermore, their geographical distribution and host specificity of the genus were thoroughly analyzed and summarized. Additionally, seven new anamorphic species of the genus Daldinia were also detected, Daldinia ehretiae sp. nov., D. jianfengensis sp. nov., D. ledongensis sp. nov., D. menghaiensis sp. nov., D. rhododendri sp. nov., D. spatholobi sp. nov., and D. thunbergiae sp. nov.

Keywords:

biodiversity; new species; morphology; phylogeny; taxonomy

1. Introduction

The genus Daldinia (Hypoxylaceae, Xylariales) was introduced by Cesati and De Notaris, as a tribute to the Swiss monk, Agostino Daldini [1], and it is now recognized as one of the largest genera within Hypoxylaceae (Xylariales, Ascomycota). The identification of Daldinia species traditionally relies on the presence of internal concentric zones beneath the perithecial layer and the detection of KOH-extractable pigments below and on the stromatal surface [2]. The current common view is that Daldinia species usually inhabited dicots. However, there have been reports of Daldinia bambusicola isolated from bamboo (monocot) in Thailand, and records of isolated Daldinia from monocot plant hosts can even be traced back to the 19th century [2,3]. Daldinia graminis and D. sacchari were reported to occur on sugarcane in India [4]. The latest world monograph of Daldinia species has employed a combination of morphological, molecular phylogenetic characteristics, and chemical classification [3]. It has also compiled morphological, structural, and chemotaxonomic data for over a thousand cultures and specimens, including preliminary phylogenetic data based on ITS sequences. Notably, several Daldinia species, including D. caldariorum, D. gelatinoides, D. loculata, D. loculatoides, and D. vernicosa, have been documented to exhibit stromata production on wood after fire events [3]. In order to determine the phylogenetic relationships of Daldinia and its allied genera, the study conducted by Wendt et al. incorporated various species from different genera into a multi-locus phylogenetic analysis, revealing that Daldinia and its relatives form a distinct branch within the Hypoxylaceae, separate from Hypoxylon and Pyrenopolyporus, and Daranagama et al. presented a comprehensive analysis and introduction of the phylogeny for key taxa within Xylariaceae, accompanied by updated illustrations and descriptions of all taxa [5,6]. Wongkanoun et al. unveiled a new record and a new species of the genus Daldinia in northern Thailand by employing traditional morphological and a multi-locus phylogenetic analysis of ITS, LSU, tub2, and rpb2 [7]. It is worth mentioning that Pažoutová et al. identified a new insect-associated, endophytic species, Daldinia hawksworthii, based on molecular data of anamorphic species and structural characteristics of their conidiophores, conidiogenous cells, and conidia, which provided a strong reference for our research [8].

China boasts abundant microbial resources across its vast territory, particularly in the southern regions characterized by intricate topography, diverse vegetation, and favorable climatic conditions. In this investigation, an extensive sampling effort was conducted across Yunnan, Hainan, Sichuan, Fujian, and Guizhou provinces of China to study fungal resources comprehensively. At present, there are few relevant articles on the systematic study of the genus Daldinia in China [3]. Therefore, the primary objective of this study was to comprehensively analyze the geographical distribution and host species exhibited by the genus Daldinia in southern China, as well as to provide detailed descriptions and explanations regarding the morphological and phylogenetic analysis of the anamorphic species of Daldinia encountered in this investigation.

2. Materials and Methods

2.1. Sample Treatment and Morphological Characterizations

Between 2022 and 2023, more than 6000 samples were collected from diseased and decayed leaves in densely vegetated areas such as forests and mountains in the Yunnan, Hainan, Sichuan, Guizhou, and Fujian provinces of China, based on the principle of collecting samples from different locations and plants as far as possible. Cut 3 × 3 mm small square leaves from the fungal infection site of the diseased and decayed leaves (the location selected for cutting is marked with a black circle and arrow in subsequent figure), disinfect the surface in 75% alcohol for 30 s, rinse once with sterile water, immerse in a 5% sodium hypochlorite solution for disinfection for 1 min, and finally rinse three times with sterile water. After disinfection, the small square leaves were dried on sterile filter paper and then transferred to the surface of Potato Dextrose Agar (PDA) (200 g potato, 20 g agar powder, 20 g dextrose, 1000 mL distilled water, and pH adjusted to 7.0) and Oatmeal Agar (OA) (25 g oats, 20 g agar powder, 1000 mL distilled water, and pH adjusted to 7.0). Three to five small square leaves can be placed on each medium; the strain number and date were marked on the medium and cultured in a constant temperature incubator at 25 °C, and the fungal growth was regularly observed. When there were more mycelia around the small square leaves, 5 × 5 mm agar blocks with mycelia were cut down with the inoculation needle and inoculated on the new PDA and OA medium and then placed in the incubator for culture. A digital camera (Canon Powershot G7X; Canon (China) Co., Ltd., Beijing, China) was utilized to capture mycelia growing on PDA medium and OA medium on day 7, day 14, and day 31, respectively. In addition, the color formed by the colonies during the cultivation process was recorded using the Pantone Colour Chart (https://www.pantone-colours.com/) (accessed on 20 June 2024). The asexual morphology of Daldinia was observed and recorded using a stereomicroscope (Olympus SZX10) and microscope (Olympus BX53). Additionally, a high-definition color digital camera (Olympus DP80) was employed to photograph the conidiogenous cells, conidia, and other structures. The microstructure of each strain (20–30 measurements per strain) was measured using the Digimizer v.5.4.7 (https://www.digimizer.com/) (accessed on 20 June 2024). All strains were stored in sterile 10% glycerol at 4 °C. Ex-type cultures were stored in the Shandong Agricultural University Culture Collection (SAUCC). Furthermore, voucher specimens were dried in the oven, sealed, and stored in the Herbarium Mycologicum Academiae Sinicae, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China (HMAS), as well as in the Herbarium of the Department of Plant Pathology, Shandong Agricultural University, Taian, China (HSAUP). Taxonomic information on all new species of the genus Daldinia has been uploaded to MycoBank (http://www.mycobank.org/) (accessed on 20 June 2024).

2.2. DNA Extraction and Sequencing

The mycelium on PDA medium was subjected to the CTAB method and BeaverBeads Plant DNA Kit (Cat. No.: 70409-20; BEAVER Biomedical Engineering Co., Ltd., Suzhou, China) for genomic DNA extraction of each of the strains [9,10,11]. PCR amplification of the genomic DNA was performed using ITSs as primers. The reaction mixture (25 μL) consisted of 2 × Hieff Canace ^® Plus PCR Master Mix (Shanghai, China) (with dye) (Yeasen Bio-technology, Shanghai, China, Cat No. 10154ES03) (12 μL), 10 μmol/μL forward primer (1 μL), 10 μmol/μL reverse primer (1 μL), 10–20 μmol/μL genomic DNA (1 μL), and distilled deionized water (10 μL). The amplified products were detected on 2% agarose electrophoresis gel and the amplification effect was examined under UV light [12]. After the gel strips with the correct band were cut off, they were recycled through the Gel Extraction Kit (Cat: AE0101-C) (Shandong Sparkjade Biotechnology Co., Ltd., Jinan, China), and the obtained DNA solutions were sent to the biological company for sequencing. Primer synthesis and DNA sequencing were conducted by Tsingke Biotechnology Co., Ltd. (Qingdao, China). The obtained sequencing results were analyzed and assembled using MEGA v.7.0 [13]. The assembled ITS sequences were BLAST searched (https://blast.ncbi.nlm.nih.gov/) (accessed on 20 June 2024) to screen out suspected Daldinia strains, and then LSU, rpb2, and tub2 primers were used to repeat the operation of the above ITS primers, and finally the sequences of ITS, LSU, rpb2, and tub2 of all suspected strains were obtained. The primer sequences and reaction conditions used in this study are presented in the Supplementary Materials [14,15,16,17]. Finally, the sequences of all strains of the Daldinia genus have been uploaded to Genbank; GenBank accession numbers are shown in Table 1.

2.3. Phylogenetic Analyses

Based on the recently published literature on the genus Daldinia, the sequences for phylogenetic analysis of the Daldinia genus were downloaded from the National Center for Biotechnology Information (https://www.ncbi.nlm.nih.gov/) (accessed on 20 June 2024) [19]. The GenBank accession numbers of all sequences used in this experiment are shown in Table 1. All sequences for phylogenetic analysis were compared and manually corrected using MEGA v.7.0 [13]. The maximum likelihood (ML), Bayesian algorithm (MB), and maximum parsimony (MP) were used to analyze all the sequence of Daldinia. The RaxML 8.2.4 of CIPRES Science Gateway V.3.3 (https://www.phylo.org/) (accessed on 20 June 2024) was used for ML trees and bootstrap analyses (MLBS), and the GTR rate parameters were optimized with the BFGS method [22,23]. The AIC of Mr Modeltest 2.2 selected the best-fit model (GTR + I + G), MrBayes 3.0B4 computed branches’ Bayesian posterior probabilities (BPP), and hierarchical likelihood ratios (hLRTs) tested results [24,25]. Three million generations in four Markov chains were ran, sampled every one hundred generations, and a burn-in value to three thousand sampled trees was set. The PAUP*4.0b10 was used for the MP analysis. All characters used for analysis were weighted equally, and gaps were missing data [26]. When MLBS ≥ 70 or BPP ≥ 0.9, it can be considered as having a good support rate [7,19]. The phylogenetic trees were viewed and adjusted with FigTree v.1.4.4 (http://tree.bio.ed.ac.uk/software/figtree) (accessed on 20 June 2024) and beautified with Adobe Illustrator CC 2019.

3. Results

3.1. Phylogenetic Analyses

The phylogenetic analysis of ITS, LSU, rpb2, and tub2 sequence data was performed to verify the interspecific relationships of Daldinia. A total of 84 sequences were used in this phylogenetic analysis, including 52 sequences of Daldinia, 12 sequences of Hypoxylon, 6 sequences of Pyrenopolyporus, 5 sequences of Annulohypoxylon, 4 sequences of Hypomontagnella, 3 sequences of Jackrogersella, and 1 sequence each of Graphostroma and Xylaria. A total of 6622 characters including gaps, 1714 of ITS, 2273 of LSU, 925 of rpb2, and 1710 of tub2 were used in this phylogenetic analysis, including 4263 constant characters, 767 variable characters that are parsimony uninformative, and 1592 parsimony informative characters. The final ML Optimization Likelihood was −56,104.543344. The trees obtained through the employment of maximum likelihood (ML) and Bayesian algorithm (MB) methods exhibit a high degree of similarity. Figure 1 shows the ML tree with the most superior score, and the corresponding maximum likelihood bootstrap support values and Bayesian posterior probabilities (MLBS/BPP) were displayed above each of the branches. This study introduced seventeen strains of the genus Daldinia for phylogenetic analysis, which were divided into 10 clades on the phylogenetic tree, representing seven new species and three known species. It is worth noting that the sequences of a large number of strains discovered during this survey were the same, belonging to three different species. Therefore, to maintain the simplicity of the phylogenetic tree, Daldinia bambusicola (SAUCC197001), D. childiae (SAUCC133401), and D. eschscholtzii (SAUCC265301) were used as representative for phylogenetic analysis. Finally, 84 strains were divided into 56 taxa on the phylogenetic tree.

3.2. Sample Information Statistics

In this investigation, a total of over 6000 samples were collected resulting in the isolation of Daldinia strains from 94 samples; we identified a total of three known species and seven new anamorphic species of the genus Daldinia; the collected information and host of each specimen are shown in Table 2, and the geographical distribution and host species are shown in Figure 2.

3.3. Taxonomy

Daldinia bambusicola Y.M. Ju, J.D. Rogers and F. San Martín, Figure 3 and Figure 4.

Description—conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores dichotomously or trichotomously branched, finely roughened, hyaline, and aseptate, with 2–3 conidiogenous cells at each terminus, 110–160 × 2.1–2.7 µm (x = 132 × 2.5 µm, n = 22). Conidiogenous cells are cylindrical, finely roughened, and hyaline, with a flattened base, bearing conidia on their apical region, 10.1–15.3 × 2.5–3.1 μm (

\bar{x}

= 12.2 × 2.7 µm, n = 24). Conidia are subglobose or ellipsoid, smooth or finely roughened, hyaline, aseptate, solitary, and produced holoblastically in sympodial sequence, 3.4–4.5 × 2.5–3.1 μm (

\bar{x}

= 3.9 × 2.8 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, the colony exhibited a diameter of 90 mm and demonstrated a growth rate of approximately 12.9 mm/day. Colonies radiated and ringed from the middle to the periphery; flat, medium dense, and aerial mycelium was more on the periphery than in the center and was white (7443); the reverse center was black (419), and the periphery was white (7443). After 7 days of cultivation on OA medium at 25 °C, the colony exhibited a diameter of 75 mm and demonstrated a growth rate of approximately 10.7 mm/day. Colonies were rough, medium dense, and aerial mycelium distributed evenly in the middle and formed irregular continuous protrusions on the periphery and was white (7541); the reverse center was dark brown (1545), and the periphery was white (7443).

Notes—based on phylogenetic analysis and spore characteristics, the strains represented by SAUCC197001 were identified as Daldinia bambusicola, as shown in Figure 1 and Figure 3 [2,3]. These strains were distributed in the Yunnan, Hainan, Sichuan, and Guizhou provinces of China, especially in warm and humid forests and mountainous areas, which belong to a tropical monsoon climate and subtropical monsoon climate, and the altitude range was 151.57 m to 1876.71 m. These strains differ from those previously reported in that they were isolated from 12 new plant hosts (Viburnum rhytidophyllum, Spatholobus suberectus, Piper nigrum, Cinnamomum verum, Koelreuteria paniculata, Ficus hirta, Schima superba, Citrus maxima, Phyllostachys heteroclada, Ageratina adenophora, Lophatherum gracile, and Ulmus pumila), as shown in Figure 4.

Figure 3. Daldinia bambusicola (type: SAUCC197001). (a) leaf of host Viburnum rhytidophyllum; (b) colony front and back after 7 days of culture on PDA; (c) colony front and back after 7 days of culture on OA; (d,e) conidiogenous cells and conidia. Scale bars: (d,e) 10 μm.

Figure 4. The host leaves of Daldinia bambusicola; relevant information is shown in Table 2. Scale bars: (a–k) 2 cm.

Daldinia childiae J.D. Rogers and Y.M. Ju, Figure 5 and Figure 6.

Description—conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores dichotomously or trichotomously branched and are smooth or finely roughened, hyaline, and aseptate, with 2–3 conidiogenous cells at each terminus, 150–220 × 2.5–3 µm (

\bar{x}

= 181 × 2.7 µm, n = 20). Conidiogenous cells are clavate, terminal enlargement, roughened, and hyaline, bearing conidia on their apical region, 14.9–32.7 × 2.8–4.4 μm (

\bar{x}

= 23.2 × 3.5 µm, n = 26). Conidia are subglobose or ellipsoid, smooth, pale yellow, hyaline, aseptate, solitary, and produced holoblastically in sympodial sequence, 5.8–9.1 × 4.1–5.8 μm (

\bar{x}

= 7.6 × 4.9 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, the colony exhibited a diameter of 90 mm and demonstrated a growth rate of approximately 12.9 mm/day. Colonies were rough, dense, and aerial mycelium distributed evenly and was white (7443); the reverse center was black (419), light brown (1395) outwards, and the colony radiated from the middle to the periphery. After 7 days of cultivation on OA medium at 25 °C, the colony exhibited a diameter of 80 mm and demonstrated a growth rate of approximately 11.4 mm/day. Colonies were rough, dense, and aerial mycelium distributed evenly and formed many punctiform protrusions and was white (7541); the reverse center was brown (1615), and the periphery was white (7541).

Notes—J.D. Rogers et al. had reported Daldinia childiae, but it was not included in the phylogenetic analysis of Daldinia in recent publications [7,19,21,28]. In this paper, the complete sequence information of ITS, LSU, rpb2, and tub2 of D. childiae was provided and included in the phylogenetic analysis. Based on sequence alignment and spore characteristics, the strains represented by SAUCC133401 were identified as D. childiae, as shown in Figure 1 and Figure 5 [3,28]. These strains were distributed in the Fujian, Yunnan, Hainan, and Sichuan provinces of China; their geographical distribution characteristics were similar to those of the D. bambusicola strains identified in this study. It is worth noting that Stadler et al. previously reported a Chinese specimen that resembles D. childiae but has an aberrant teleomorphic morphology [3]. The difference from previous reports is that the D. childiae found in this investigation was isolated from 10 new plant hosts (Machilus nanmu, Eurya japonica, Piper nigrum, Microstegium vimineum, Pseudosasa japonica, Litsea cubeba, Quercus glauca, Schima superba, Castanopsis calathiformis, and Symplocos sumuntia), as shown in Figure 6.

Figure 5. Daldinia childiae (type: SAUCC133401). (a) leaf of host Machilus nanmu; (b) colony front and back after 7 days of culture on PDA; (c) colony front and back after 7 days of culture on OA; (d,e) conidiogenous cells and conidia. Scale bars: (d,e) 10 μm.

Figure 6. The host leaves of Daldinia childiae; relevant information is shown in Table 2. Scale bars: (a–i) 2 cm.

Daldinia eschscholtzii Rehm, Figure 7 and Figure 8.

Description—conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous, dichotomously or trichotomously branched, finely roughened, hyaline, and aseptate, with 2–3 conidiogenous cells at each terminus, 120–214 × 2.3–4.1 µm (

\bar{x}

= 176 × 3.4 µm, n = 21). Conidiogenous cells are cylindrical, finely roughened, and hyaline, with a flattened base, bearing conidia on their apical region, 14.9–22.7 × 2.1–3.6 μm (

\bar{x}

= 18.3 × 2.7 µm, n = 26). Conidia are ellipsoid or dacryoid, smooth to finely roughened, hyaline, aseptate, and solitary, mostly with a flattened base and produced holoblastically in sympodial sequence, 4.9–6.8 × 2.3–3.5 μm (

\bar{x}

= 5.7 × 2.8 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, the colony exhibited a diameter of 88 mm and demonstrated a growth rate of approximately 12.6 mm/day. Colonies radiated from the middle to the periphery, rough, medium dense, aerial mycelium was more on the periphery than in the center, and formed some protrusions and was white (7443); the reverse center formed a black (419) area with irregular edges surrounded by white (7443). After 7 days of cultivation on OA medium at 25 °C, the colony exhibited a diameter of 86 mm and demonstrated a growth rate of approximately 12.2 mm/day. Colonies were rough, medium dense, and aerial mycelium distributed evenly and was white (7443); the reverse center was light brown (1395), the periphery was white (7443), and multiple rings appeared from the middle to the periphery.

Notes—based on phylogenetic analysis and spore characteristics, the strains represented by SAUCC265301 were identified as Daldinia eschscholtzii, as shown in Figure 1 and Figure 7 [3]. According to the survey, it was extensively found in the Fujian, Yunnan, Hainan, Sichuan, and Guizhou provinces of China. Particularly prevalent in warm, moist and vegetated forests, mountains, plains, and wetlands at elevations ranging from 154.85 m to 1876.7 m. This survey has discovered 58 new plant hosts of Daldinia; they belong to 38 families, mainly Lauraceae, Moraceae, and Poaceae, as shown in Figure 8.

Figure 7. Daldinia eschscholtzii (type: SAUCC265301). (a) leaf of host Lysimachia clethroides; (b) colony front and back after 7 days of culture on PDA; (c) colony front and back after 7 days of culture on OA; (d,e) conidiogenous cells and conidia. Scale bars: (d,e) 10 μm.

Figure 8. The host leaves of Daldinia eschscholtzii; relevant information is shown in Table 2. Scale bars: (a1–o) 2 cm.

Daldinia ehretiae C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 9.

MycoBank—MB853131

Etymology—the epithet “ehretiae” refers to the name of the host plant, Ehretia acuminata.

Material studied—China, Yunnan Province, Jinghong City, Sancha River, 22°10′10″ N, 100°51′49″ E, on diseased leaves of Ehretia acuminata, 19 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352914, ex-type culture SAUCC228302.

Description—conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous or dichotomously branched and are finely roughened, hyaline, and aseptate, with 1–2 conidiogenous cells at each terminus, 100–210 × 3.1–4.3 µm (

\bar{x}

= 165 × 3.6 µm, n = 23). Conidiogenous cells are cylindrical, finely roughened, and hyaline, bearing conidia on their apical region, 16.8–24.5 × 2.7–4.1 µm (

\bar{x}

= 19.3 × 3.5 µm, n = 25). Conidia are ellipsoid or cylindrical, smooth to finely roughened, hyaline, aseptate, and solitary, mostly with a flattened base, and produced holoblastically in sympodial sequence, 4.2–6.6 × 1.7–2.8 µm (

\bar{x}

= 5.4 × 2.2 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies radiated and ringed from the middle to the periphery, showing black (419), pale green (365), grey (422), and white (7541) in sequence, flat, medium dense, and aerial mycelium distributed evenly; the reverse was similar to the front. After 14 days, colonies formed multiple white (7443) rings, dense, and aerial mycelium grey (423); the reverse center and periphery was black (426), and the intermediate part was light green (373). After 31 days, colonies changed from gray to turquoise brown (403); the reverse was mostly black (426). The average growth rate on PDA medium was 11.7 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies formed a dark green (378) ring, and the rest was mostly white (7443), rough, medium dense, and aerial mycelium was more on the periphery than in the center; the reverse was similar to the front. After 14 days, colonies alternated gray (425) and white (7541) rings from the middle to the periphery, dense, and aerial mycelium distributed evenly; the reverse alternated brown (1615) and white (7443) rings from the middle to the periphery. After 31 days, colonies changed from gray to light gray (427); the reverse was mostly brown (1615). The average growth rate on OA medium was 11.6 mm/day.

Additional material studied—China, Yunnan Province, Jinghong City, Sancha River, 22°10′10″ N, 100°51′49″ E, on diseased leaves of Ehretia acuminata, 19 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 228303, living culture SAUCC228303.

Notes—the phylogenetic analysis of ITS, LSU, rpb2, and tub2 sequences revealed a close relationship between Daldinia ehretiae and another newly discovered species, D. thunbergiae. Daldinia ehretiae is different from D. thunbergiae by 69/559 bp in ITS, 17/1259 bp in LSU, and 201/1393 bp in tub2. Morphologically, D. ehretiae and D. thunbergiae differ in that there are a fewer number of conidiogenous cells (1–2 × 1–3) at the terminus of conidiophores and larger conidiogenous cells (16.8–24.5 × 2.7–4.1 vs. 6.7–17.3 × 1.9–2.5 µm). In addition, there is little difference in conidia size between D. ehretiae and D. thunbergiae, but the conidial shape of D. ehretiae is mostly ellipsoid or cylindrical, and the conidia of D. thunbergiae are mostly teardrop shaped except ellipsoid. Therefore, D. ehretiae can be identified as a new anamorphic species of Daldinia via phylogenetic and morphological comparison.

Figure 9. Daldinia ehretiae (holotype: HMAS352914). (a) leaf of host plant Ehretia acuminata; (b–d) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (e–g) colony front and back after 7 days, 14 days, and 31 days culture on OA; (h–j) conidiogenous cells and conidia. Scale bars: (h–j) 10 μm.

Daldinia rhododendri C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 10.

MycoBank—MB853135

Etymology—the epithet “rhododendri” refers to the name of the host plant, Rhododendron decorum.

Material studied—China, Yunnan Province, Diqing Prefecture, Shangri-la City, 27°58′43″ N, 99°34′24″ E, on diseased leaves of Rhododendron decorum, 28 June 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352918, ex-type culture SAUCC460001.

Description—conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous or dichotomously branched and are rare, smooth or finely roughened, hyaline, and septate, with conidiogenous cells at each terminus, 40–90 × 1.4–2.0 µm (

\bar{x}

= 74 × 1.7 µm, n = 21). Conidiogenous cells are cylindrical or ampuliform, rare, smooth to finely roughened, and hyaline, bearing conidia on their apical region, 5.9–11.6 × 1.1–2.9 µm (

\bar{x}

= 8.6 × 2.3 µm, n = 26). Conidia are ellipsoid, cylindrical or banana shaped, smooth, hyaline, aseptate, solitary, and produced from percurrently proliferating conidiogenous cells, 3.2–5.1 × 1.1–2.3 µm (

\bar{x}

= 4.1 × 1.7 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies were rough, dense, and aerial mycelium distributed evenly and was white (7443) and grey (424) from the middle out; the reverse center was white (7443), with irregular areas of reddish brown (469) around. After 14 days, colonies formed a few white (7541) fluffy balls and a circle of white (7541) fluffy mycelium around; aerial mycelium was dark grey (425); the reverse center was white (7443), with irregular areas of reddish brown (469) and pale green (384) around. After 31 days, colonies had not changed significantly; the reverse was predominantly brown (1405) and black (426). The average growth rate on PDA medium was 12 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies alternated dark gray (425) and white (7443) with rings from the middle to the periphery and was rough, medium dense, with aerial mycelium distributed evenly; the reverse was white (7443). After 14 days, colonies formed a few white (7541) fluffy balls, dense; the reverse exhibited an irregular dark gray (425) ring. After 31 days, colonies changed from dark gray to gray (423); the reverse exhibited some irregular yellow green (582) areas. The average growth rate on OA medium was 11.1 mm/day.

Additional material studied—China, Yunnan Province, Diqing Prefecture, Shangri-la City, 27°58′43″ N, 99°34′24″ E, on diseased leaves of Rhododendron decorum, 28 June 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 460002, living culture SAUCC460002.

Notes—the phylogenetic analysis of ITS, LSU, rpb2, and tub2 sequences revealed a close relationship between Daldinia rhododendri, D. pyrenaica, and D. childiae, and they are both on a branch with better support (MLBS/BPP = 100/1). Daldinia rhododendri is different from D. pyrenaica by 21/552 bp in ITS, 4/982 bp in LSU, and 29/1342 bp in tub2 and different from D. childiae by 20/552 bp in ITS, 6/982 bp in LSU, and 18/1368 bp in tub2. Morphologically, D. rhododendri and D. pyrenaica differ obviously in conidiogenous cells (5.9–11.6 × 1.1–2.9 vs. 10.0–25.0 × 2.5–3.0 µm) and conidia (3.2–5.1 × 1.1–2.3 vs. 6.5–7.0 × 4.0–5.0 μm) [3,29]. Daldinia rhododendri and D. childiae also differ obviously in conidiogenous cells (5.9–11.6 × 1.1–2.9 vs. 10.0–25.0 × 3.0–4.0 µm) and conidia (3.2–5.1 × 1.1–2.3 vs. 7.0–9.0 × 4.5–5.5 μm) [3,28]. Therefore, D. rhododendri can be identified as a new anamorphic species of Daldinia by phylogenetic and morphological comparison.

Figure 10. Daldinia rhododendri (holotype: HMAS352918). (a) leaf of host plant Rhododen dron decorum; (b–d) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (e–g) colony front and back after 7 days, 14 days, and 31 days culture on OA; (h–j) conidiogenous cells and conidia. Scale bars: (h–j) 10 μm.

Daldinia spatholobi C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 11.

MycoBank—MB853136

Etymology—the epithet “spatholobi” refers to the name of the host plant, Spatholobus suberectus.

Material studied—China, Yunnan Province, Jinghong City, Xishuangbanna primitive Forest Park, 22°1′52″ N, 100°52′36″ E, on diseased leaves of Spatholobus suberectus, 17 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352919, ex-type culture SAUCC203501.

Description—Conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous, dichotomously, or trichotomously branched and are roughened, hyaline, and septate, with 1–3 conidiogenous cells at each terminus, 90–150 × 2.9–4.1 µm (

\bar{x}

= 135 × 3.4 µm, n = 22). Conidiogenous cells are cylindrical or clavate, roughened, and hyaline, with a flattened base, bearing conidia on their apical region, 9.5–14.9 × 2.4–3.0 µm (

\bar{x}

= 12.3 × 2.6 µm, n = 25). Conidia are ellipsoid or teardrop shaped, smooth to finely roughened, hyaline, aseptate, solitary, and produced holoblastically in sympodial sequence, 4.3–6.1 × 2.6–3.7 µm (

\bar{x}

= 5.6 × 3.4 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies alternated pale brown (1245), white (7443), pale green (390), and white (7443) with rings from the middle to the periphery and were flat, medium dense, and with aerial mycelium more in the center than on the periphery; the reverse was similar to the front, but the center was black (419). After 14 days, colonies formed irregular green (377) areas and was dense with aerial mycelium distributed evenly; the reverse center was black (419), brown (1255) with irregular area outside, and white (7443) outwards. After 31 days, colonies were white (7527) in the center, and the rest was mostly black (419); the reverse was also mostly brown (1405). The average growth rate on PDA medium was 10.6 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies formed a black (426) circular area in the center and was white (7443) outwards and flat, medium dense, and with aerial mycelium distributed evenly; the reverse center was brown (1405) and mostly white (7443) on the outside. After 14 days, colonies ringed from the middle to the periphery, showing black (426), pale brown (1395), grey (423), dark green (371), and white (7541) colors and were dense; the reverse center was black (419) and formed a regular brown (1405) ring. After 31 days, colonies darkened in color; the reverse has not changed significantly. The average growth rate on OA medium was 9.7 mm/day.

Additional material studied—China, Yunnan Province, Jinghong City, Xishuangbanna primitive Forest Park, 22°1′52″ N, 100°52′36″ E, on diseased leaves of Spatholobus suberectus, 17 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 203502, living culture SAUCC203502.

Notes—the phylogenetic analysis of ITS, LSU, rpb2, and tub2 sequences revealed a close relationship between Daldinia spatholobi and D. kretzschmarioides, and they are both on a branch with better support (MLBS/BPP = 99/1). Daldinia spatholobi was different from D. kretzschmarioides by 6/961 bp in LSU and 25/1066 bp in tub2. Morphologically, compared to D. kretzschmarioides, D. spatholobi has shorter conidiogenous cells (2.4–3.0 vs. 3.0–4.0 µm) and similar conidia sizes. But the conidia of D. spatholobi are teardrop shaped in addition to ellipsoid, with a finely roughened surface. Under the same conditions, D. spatholobi exhibits regular circular pigment precipitation on OA medium with fewer mycelium, while D. kretzschmarioides exhibits irregular pigment precipitation with more mycelium [7]. Therefore, D. spatholobi can be identified as a new anamorphic species of Daldinia by phylogenetic and morphological comparison.

Figure 11. Daldinia spatholobi (holotype: HMAS352919). (a) leaf of host plant Spatholobus suberectus; (b–d) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (e–g) colony front and back after 7 days, 14 days, and 31 days culture on OA; (h–j) conidiogenous cells and conidia. Scale bars: (h–j) 10 μm.

Daldinia thunbergiae C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 12.

MycoBank—MB853137

Etymology—the epithet “thunbergiae” refers to the name of the host plant, Thunbergia grandiflora.

Material studied—China, Yunnan Province, Jinghong City, Sancha River, 22°10′10″ N, 100°51′49″ E, on diseased leaves of Thunbergia grandiflora, 19 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352920, ex-type culture SAUCC228601.

Description—Conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous, dichotomously, or trichotomously branched and are roughened, hyaline, and septate, with 1–4 conidiogenous cells at each terminus, 70–220 × 1.8–4.1 µm (

\bar{x}

= 150 × 3.3 µm, n = 21). Conidiogenous cells are cylindrical or clavate, finely roughened, and hyaline, bearing conidia on their apical region, 6.7–17.3 × 1.9–2.5 µm (

\bar{x}

= 11.5 × 2.2 µm, n = 26). Conidia are ellipsoid or teardrop shaped, finely roughened, hyaline, aseptate, solitary, mostly with a flattened base, and produced holoblastically in sympodial sequence, 3.2–5.0 × 2.2–3.1 µm (

\bar{x}

= 4.2 × 2.7 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies radiated from the middle to the periphery, showing white (7443), dark green (385), and white (7443) colors and were rough, medium dense, and with aerial mycelium more in the center than on the periphery; the reverse center was dark green (385), with an incomplete dark green (385) ring outwards, and the rest was white (7443). After 14 days, colonies were mostly dark gray (425), plat, dense, and with aerial mycelium distributed evenly; the reverse center was mostly black (426), surrounded by many black (426) spots and irregular pale green (391) areas, and the rest was white (7443). After 31 days, colonies changed from dark gray to dark brown (1545); white (7541) mycelium appeared in the center; the reverse periphery was mostly brown (1615). The average growth rate on PDA medium was 11.1 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies formed multiple white (7541) rings and the center was black (419), plat, and medium dense, with aerial mycelium distributed evenly; the reverse was similar to the front, but the center was brown (1615). After 14 days, colonies formed a white (7443) ring in the outermost ring and the rest was mostly gray (424) and dense; the reverse was similar to the front, but the center and some irregular parts were dark green (378). After 31 days, colonies changed from gray to light brown (1255); the reverse was also mostly brown (1265). The average growth rate on OA medium was 11.3 mm/day.

Additional material studied—China, Yunnan Province, Jinghong City, Sancha River, 22°10′10″ N, 100°51′49″ E, on diseased leaves of Thunbergia grandiflora, 19 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 228602, living culture SAUCC228602.

Notes—Daldinia thunbergiae is closely related to another newly discovered species in this study, D. ehretiae, on the phylogenetic tree of Daldinia. Based on the differences in sequence, conidiophores, conidiogenous cells, and conidia between the two species mentioned above, D. thunbergiae can be identified as a new anamorphic species of Daldinia.

Figure 12. Daldinia thunbergiae (holotype: HMAS352920). (a) leaf of host plant Thunbergia grandiflora; (b–d) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (e–g) colony front and back after 7 days, 14 days, and 31 days culture on OA; (h–j) conidiogenous cells and conidia. Scale bars: (h–j) 10 μm.

Daldinia jianfengensis C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 13.

MycoBank—MB853132

Etymology—the epithet “jianfengensis” refers to the name of the strain location, Jianfeng Town.

Material studied—China, Hainan Province, Ledong County, Jianfeng Town, 18°41′19″ N, 108°51′31″ E, on decayed leaves, 12 April 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352915, ex-type culture SAUCC373804.

Description—conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous, dichotomously, or trichotomously branched and are finely roughened, hyaline, and septate, with 1–4 conidiogenous cells at each terminus, 70–120 × 2.9–4.4 µm (

\bar{x}

= 97 × 3.6 µm, n = 22). Conidiogenous cells are cylindrical, finely roughened, and hyaline, bearing conidia on their apical region, 12.1–16.9 × 2.6–3.6 µm (

\bar{x}

= 15.1 × 3.2 µm, n = 25). Conidia are subglobose or ellipsoid, smooth to finely roughened, hyaline, aseptate, solitary, mostly with a flattened base, and produced holoblastically in sympodial sequence, 3.2–5.5 × 2.6–3.7 µm (

\bar{x}

= 4.3 × 3.5 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies exhibited alternated white (7443) and gray (422) rings from the middle to the periphery, with periphery irregularity, flat, and medium dense; the reverse center was black (426), dark green (385) irregular area outside, and white (7443) outwards. After 14 days, colonies formed a round, white (7443) fluff-like area in the center, dense, and aerial mycelium distributed evenly, grey (424); the reverse exhibited alternating black (419) and white (7443) irregular rings from the middle to the periphery. After 31 days, colonies and reverse had not changed significantly. The average growth rate on PDA medium was 11.6 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies were black (419), gray (425), and white (7443) from the middle out, flat, and medium dense; the reverse center was gray (424), with a circle of black (419) on the outside and mostly white (7541) on the outside. After 14 days, colonies alternated gray (424) and white (7443) from the middle to the periphery, dense, and aerial mycelium distributed evenly; the reverse alternated gray (423) and white (7443) with regular rings from the middle to the periphery. After 31 days, colonies changed from gray to pale brown (1265); the reverse was slightly darker. The average growth rate on OA medium was 11.4 mm/day.

Additional material studied—China, Hainan Province, Ledong County, Jianfeng Town, 18°41′19″ N, 108°51′31″ E, on decayed leaves, 12 April 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 373805, living culture SAUCC373805.

Notes—The phylogenetic analysis of ITS, LSU, rpb2, and tub2 sequences revealed a close relationship between Daldinia jianfengensis and another newly discovered species, D. ledongensis, and they are both on a branch with better support (MLBS/BPP = 100/1). D. jianfengensis was different from D. ledongensis by 56/549 bp in ITS, 12/1277 bp in LSU, and 5/1038 bp in tub2. Morphologically, the difference between D. jianfengensis and D. ledongensis is that there are more conidiogenous cells (1–3 × 1–2) at the terminus of conidiophores and conidiogenous cells are generally long (12.1–16.9 vs. 8.6–15.1 µm) and are mostly distributed at the terminus of conidiophores. In addition, the conidia of D. jianfengensis are wider (2.6–3.7 vs. 1.4–2.0 μm) and produce more conidia under the same conditions. The conidial shape of D. jianfengensis is subglobose or ellipsoid, which is different from that of the conidia of D. ledongensis with an ellipsoid or fusiform shape. It is worth noting that under the same culture conditions, whether in OA medium or PDA medium, the medium of D. jianfengensis will show circular dark pigmentation, and the mycelium will be darker, while the medium of D. ledongensis only has a small part of pigmentation and the mycelium was always white. Therefore, D. jianfengens can be identified as a new anamorphic species of Daldinia by phylogenetic and morphological comparison.

Figure 13. Daldinia jianfengensis (holotype: HMAS352915). (a–c) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (d–f) colony front and back after 7 days, 14 days, and 31 days culture on OA; (g–i) conidiogenous cells and conidia. Scale bars: (g–i) 10 μm.

Daldinia ledongensis C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 14.

MycoBank—MB853133

Etymology—the epithet “ledongensis” refers to the name of the strain location, Ledong County.

Material studied—China, Hainan Province, Ledong County, 18°41′18″ N, 108°51′31″ E, on decayed leaves, 12 April 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352916, ex-type culture SAUCC393602.

Description—Conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores mononematous or dichotomously branched and are rare, smooth or finely roughened, hyaline, and aseptate, with a conidiogenous cells at each terminus, 120–200 × 1.7–2.1 µm (

\bar{x}

= 157 × 1.8 µm, n = 21). Conidiogenous cells are clavate, rare, smooth to finely roughened, and hyaline, with a flattened base, bearing conidia on their apical region, 8.6–15.1 × 1.2–3.4 µm (

\bar{x}

= 11.3 × 2.4 µm, n = 24). Conidia are ellipsoid or fusiform, smooth to finely roughened, hyaline, aseptate, solitary, and produced from percurrently proliferating conidiogenous cells, 3.2–4.0 × 1.4–2.0 µm (

\bar{x}

= 3.5 × 1.6 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies radiated from the middle to the periphery, rough, medium dense, with aerial mycelium more on the periphery than in the center and was white (7443); the reverse was similar to the front, but the center was hazel (471). After 14 days, colonies were dense, and aerial mycelium distributed evenly; the reverse was similar to the front, but the center was brown (1405). After 31 days, colonies became denser; the reverse had not changed significantly. The average growth rate on PDA medium was 10.7 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies were rough, medium dense, and aerial mycelium was more on the periphery than in the center and was white (7443); the reverse center was brown (1615), mostly white (7443) on the outside. After 14 days, colonies were dense; the reverse exhibited some dark brown (1545) areas. After 31 days, colonies became denser; the reverse exhibited many irregular brown (1395) areas. The average growth rate on OA medium was 11.4 mm/day.

Additional material studied—China, Hainan Province, Ledong County, 18°41′18″ N, 108°51′31″ E, on decayed leaves, 12 April 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 393603, living culture SAUCC393603.

Notes—Daldinia ledongensis was closely related to another newly discovered species, D. jianfengensis, on the phylogenetic tree of Daldinia. Based on the differences in sequence, conidiophores, conidiogenous cells, conidia, and medium between the two species mentioned above, D. ledongensis can be identified as a new anamorphic species of Daldinia.

Figure 14. Daldinia ledongensis (holotype: HMAS352916). (a–c) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (d–f) colony front and back after 7 days, 14 days, and 31 days culture on OA; (g,h) conidiogenous cells and conidia. Scale bars: (g,h) 10 μm.

Daldinia menghaiensis C.Z. Yin, Z.X. Zhang and X.G. Zhang, sp. nov. Figure 15.

MycoBank—MB853134

Etymology—the epithet “menghaiensis” refers to the name of the strain location, Menghai County.

Material studied—China, Yunnan Province, Menghai County, 21°55′25″ N, 100°35′41″ E, on decayed leaves, 18 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, holotype HMAS352917, ex-type culture SAUCC242404.

Description—Conidiophores exhibit a virgariella-like to nodulisporium-like branching pattern [27]. Conidiophores dichotomously or trichotomously branched, occasionally branched from the conidiogenous region, and are finely roughened, hyaline, and septate, with 1–2 conidiogenous cells at each terminus, 80–150 × 1.9–3.4 µm (

\bar{x}

= 115 × 2.6 µm, n = 22). Conidiogenous cells are cylindrical or clavate, finely roughened, and hyaline, bearing conidia on their apical region, 16.9–23.5 × 2.0–3.5 µm (

\bar{x}

= 19.3 × 2.8 µm, n = 27). Conidia are ellipsoid, subglobose or dacryoid, roughened, hyaline, aseptate, solitary, mostly with flattened base, and produced holoblastically in sympodial sequence, 4.7–8.2 × 3.1–4.0 µm (

\bar{x}

= 5.7 × 3.5 µm, n = 30). The teleomorph was not discovered.

Culture characteristics—after 7 days of cultivation on PDA medium at 25 °C, colonies radiated and ringed from the middle to the periphery, flat, medium dense, and with aerial mycelium more on the periphery than in the center and was white (7443); the reverse was similar to the front, but the center was pale brown (1375). After 14 days, colonies were dense; the reverse center was brown (1395), there appeared some light green (382) areas. After 31 days, colonies became rough; the reverse center was black (419), and the shape of the colonies became irregular. The average growth rate on PDA medium was 12.8 mm/day. After 7 days of cultivation on OA medium at 25 °C, colonies were flat, medium dense, and aerial mycelium was rare in the central circular region and was white (7443); the reverse was similar to the front. After 14 days, colonies formed many dark green (378) spots, dense, with aerial mycelium distributed evenly; the reverse was similar to the front. After 31 days, colonies changed from white to gray (424); the reverse exhibited many small irregular brown (1615) areas. The average growth rate on OA medium was 10.3 mm/day.

Additional material studied—China, Yunnan Province, Menghai County, 21°55′25″ N, 100°35′41″ E, on decayed leaves, 18 March 2023, C.Z. Yin, Z.X. Zhang and X.G. Zhang, HSAUP 242405, living culture SAUCC242405.

Notes—the phylogenetic analysis of ITS, LSU, rpb2, and tub2 sequences revealed a close relationship between Daldinia menghaiensis, D. subvernicosa, and D. vernicosa, and they are both on a branch with better support (MLBS/BPP = 99/1). Daldinia menghaiensis was different from D. subvernicosa by 25/464 bp in ITS, 16/1107 bp in LSU, 39/684 bp in rpb2, and 57/949 bp in tub2 and different from D. vernicosa by 35/552 bp in ITS, 19/1226 bp in LSU, 54/867 bp in rpb2, and 62/1080 bp in tub2. Morphologically, compared with Daldinia vernicosa, the number of conidiogenous cells at the terminus of conidiophores in D. menghaiensis is less (1–2 vs. 1–4) and is generally long (16.9–23.5 vs. 8.0–23.0 µm), but the conidia are narrower (3.1–4.0 vs. 4.5–6.0 µm) [3]. Under the same conditions, the medium of D. menghaiensis always produces pigmentation, and mycelia become darker, but D. subvernicosa does not [7]. Therefore, D. menghaiensis can be identified as a new anamorphic species of Daldinia by phylogenetic and morphological comparison.

Figure 15. Daldinia menghaiensis (holotype: HMAS352917). (a–c) colony front and back after 7 days, 14 days, and 31 days culture on PDA; (d–f) colony front and back after 7 days, 14 days, and 31 days culture on OA; (g,h) conidiogenous cells and conidia. Scale bars: (g,h) 10 μm.

4. Discussion

This study investigated the fungal resources in five provinces in southern China, and the strains of Daldinia isolated in this survey were mainly collected from forests and mountainous areas, with altitudes ranging from 151.57 to 1876.7 m.a.s.l. These strains were found in all five provinces of southern China, which proved that they were also widely distributed in southern China. The number and types of strains were the highest in Yunnan Province, followed by Hainan Province, and the least were in Guizhou Province. The overall distribution of all strains in each province is shown in Figure 2a. We noticed that with similar sample sizes collected in each province, more strains of Daldinia were collected in the Yunnan and Hainan provinces in the tropical monsoon climate zone than in the other three provinces in the subtropical monsoon climate zone. Our preliminary hypothesis posits that these primary forests and mountains, boasting warm and humid climate conditions and abundant vegetation, may harbor more Daldinia as well as other fungal. The plant specimens collected in this survey were divided into diseased and decayed leaves, and 84 new host plants were determined by identifying the strains of Daldinia. They belong to 43 plant families, among which Lauraceae (10.84%), Poaceae (9.64%), Moraceae (6.02%), Theaceae (6.02%), and Fagaceae (4.82%) have a relatively high proportion, and there is little difference in the proportion of other families. The number of each plant family is shown in Figure 2b. In addition, dicots accounted for 84.34% of all plant hosts, while monocots were only Poaceae, Zingiberaceae, Iridaceae, and Asparagaceae. Therefore, it can be preliminarily inferred that the genus Daldinia does not exhibit strong specificity of host, and dicots account for the majority of hosts.

Since the establishment of the genus Daldinia, continuous advancements have been made in the methods utilized for its identification. Now, a widely accepted approach for identifying Daldinia involves phylogenetic analysis based on ITS, LSU, rpb2, and tub2 combined with morphological comparison. A total of 104 records encompassing various species of Daldinia were retrieved from the Index Fungorum (https://www.indexfungorum.org/, accessed on 7 September 2024), among which most of the species with complete ITS, LSU, rpb2, and tub2 sequences are included in Table 1. We also used this method to identify three known species (Daldinia bambusicola, D. childiae, and D. eschscholtzii) and seven new anamorphic species (D. ehretiae, D. jianfengensis, D. ledongensis, D. menghaiensis, D. rhododendri, D. spatholobi, and D. thunbergiae). In this study, we not only provided sequence information for new species but also perfected sequence data for known species, such as the rpb2 and tub2 sequence data of D. childiae, after species were confirmed through anamorph characteristics and multiple sequence alignments. The Daldinia strains were cultured using PDA and OA mediums, and the colony morphology at 7 days, 14 days, and 31 days of culture was observed and recorded, which could also assist in the identification of the strain while showing the colony changes. In the process of cultivating the strains of Daldinia, we also noticed an interesting phenomenon: taking the new species discovered this time as an example, whether on PDA medium or OA medium, Daldinia can be divided into two categories. One type produces a large number of spores and forms a regular large-area pigment precipitation area on the medium, and most of them form circular pigment precipitation around 2 weeks, such as D. ehretiae, D. jianfengensis, D. menghaiensis, D. rhododendri, D. spatholobi, and D. thunbergiae. And another type features the production of very small amounts of spores and the formation of small areas of pigment precipitation on the culture medium, such as D. ledongensis. Furthermore, it is worth noting that there have been numerous reports that Daldinia are endophytes, some even related to insects, and spores of Daldinia typically exhibit both teleomorph and anamorph modes, but the specimens obtained during this survey were found on diseased leaves and did not directly produce spores of teleomorph in their natural habitat. So, inducing Daldinia to produce spores of teleomorph in the laboratory presents an intriguing yet challenging task. At the same time, we will continue to conduct field collection work on teleomorphic species of Daldinia in the next step and improve the teleomorphic data of Daldinia found in this survey as soon as possible. Interestingly, due to the abundance of secondary metabolites within the genus Daldinia, there has been a growing trend towards utilizing HPLC profiling for metabolite analysis and incorporating chemical classification into the identification process. In their investigation of Xylariales in Thailand, Wongkanoun et al. employed morphological characters, phylogenetic analysis based on multi-locus sequences, and comprehensive analysis of secondary metabolites based on high-performance liquid chromatography-diode array detection and mass spectrometry, leading to the identification of three novel species, D. flavogranulata, D. phadaengensis, and D. chiangdaoensis [19]. Moving forward, we will continue conducting comprehensive research on resource exploration and metabolic characteristic analysis of the genus Daldinia.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/jof10100700/s1, Table S1: PCR primer sequence and reaction conditions used in this experiment.

Author Contributions

Conceptualization, C.Y.; methodology, Z.Z.; software, C.Y.; validation, S.W. and Z.Z.; formal analysis, C.Y.; investigation, W.L.; resources, C.Y. and W.L.; data curation, C.Y. and W.L.; writing—original draft preparation, C.Y.; writing—review and editing, C.Y.; visualization, Z.Z.; supervision, X.Z.; project administration, X.Z.; funding acquisition, X.Z. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the National Natural Science Foundation of China (nos. 32170012, 32300011, 32470004, 31400019), Ji’nan City’s ‘New University 20 Policies’ Initiative for Innovative Research Teams Project (202228028), Innovative Agricultural Application Technology Project of Jinan City (CX202210), and Key Technological Innovation Program of Shandong Province, China (2022CXGC020710).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

All datasets in this study are included in this article/Supplementary Materials.

Conflicts of Interest

The authors declare no conflicts of interest.

References

Cesati, V.; De Notaris, G. Schema di classificazione degle sferiacei italici aschigeri piu’ o meno appartenenti al genere Sphaeria nell’antico significato attribuitoglide Persono. Comment. Della Soc. Crittogamologica Ital. 1 1863, 4, 177–420. [Google Scholar]
Ju, Y.M.; Rogers, J.D.; San Martín, F. A revision of the genus Daldinia. Mycotaxon 1997, 61, 243–293. [Google Scholar]
Stadler, M.; Læssøe, T.; Fournier, J.; Decock, C.; Schmieschek, B.; Tichy, H.V.; Peršoh, D. A polyphasic taxonomy of Daldinia (Xylariaceae). Stud. Mycol. 2014, 77, 1–143. [Google Scholar] [CrossRef]
Dargan, J.S.; Thind, K.S. Xylariaceae of India. VIII Genus Daldinia Ces & de Not-a further segregation into two new subgenera. Kavaka 1985, 12, 113–118. [Google Scholar]
Wendt, L.; Sir, E.B.; Kuhnert, E.; Heitkämper, S.; Lambert, C.; Hladki, A.I.; Romero, A.I.; Luangsa-ard, J.J.; Srikitikulchai, P.; Peršoh, D.; et al. Resurrection and emendation of the Hypoxylaceae, recognised from a multi-gene genealogy of the Xylariales. Mycol. Prog. 2018, 17, 115–154. [Google Scholar] [CrossRef]
Daranagama, D.A.; Hyde, K.D.; Sir, E.B.; Thambugala, K.M.; Tian, Q.; Samarakoon, M.C.; McKenzie, E.H.C.; Jayasiri, S.C.; Tibpromma, S.; Bhat, J.D.; et al. Towards a natural classification and backbone tree for Graphostromataceae, Hypoxylaceae, Lopadostomataceae and Xylariaceae. Fungal Divers. 2018, 88, 1–165. [Google Scholar] [CrossRef]
Wongkanoun, S.; Wendt, L.; Stadler, M.; Luangsa-ard, J.J.; Srikitikulchai, P. A novel species and a new combination of Daldinia from Ban Hua Thung community forest in the northern part of Thailand. Mycol. Prog. 2019, 18, 553–564. [Google Scholar] [CrossRef]
Pažoutová, S.; Follert, S.; Bitzer, J.; Keck, M.; Surup, F.; Šrůtka, P.; Holuša, J.; Stadler, M. A new endophytic insect-associated Daldinia species, recognised from a comparison of secondary metabolite profiles and molecular phylogeny. Fungal Divers. 2013, 60, 107–123. [Google Scholar] [CrossRef]
Doyle, J.J. Isolation of plant DNA from fresh tissue. Focus 1990, 12, 13–15. [Google Scholar]
Guo, L.D.; Hyde, K.D.; Liew, E.C.Y. Identification of endophytic fungi from Livistona chinensis based on morphology and rDNA sequences. New Phytol. 2000, 147, 617–630. [Google Scholar] [CrossRef]
Wang, S.; Liu, X.; Xiong, C.; Gao, S.; Xu, W.; Zhao, L.; Song, C.; Liu, X.; James, T.Y.; Li, Z.; et al. ASF1 regulates asexual and sexual reproduction in Stemphylium eturmiunum by DJ-1 stimulation of the PI3K/AKT signaling pathway. Fungal Divers. 2023, 123, 159–176. [Google Scholar] [CrossRef]
Zhang, Z.X.; Liu, R.Y.; Liu, S.B.; Mu, T.C.; Zhang, X.G.; Xia, J.W. Morphological and phylogenetic analyses reveal two new species of Sporocadaceae from Hainan, China. MycoKeys 2023, 88, 171–192. [Google Scholar] [CrossRef] [PubMed]
Kumar, S.; Stecher, G.; Tamura, K. MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets. Mol. Biol. Evolut. 2016, 33, 1870–1874. [Google Scholar] [CrossRef] [PubMed]
White, T.J.; Bruns, L.; Lee, S.; Taylor, J. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. PCR Protoc. A Guide Methods Appl. 1990, 38, 315–322. [Google Scholar] [CrossRef]
Bunyard, B.A.; Nicholson, M.S.; Royse, D.J. A systematic assessment of Morchella using RFLP analysis of the 28S ribosomal RNA gene. Mycologia 1994, 86, 762–772. [Google Scholar] [CrossRef]
Liu, Y.L.; Whelen, S.; Hall, B.D. Phylogenetic relationships among ascomycetes: Evidence from and RNA polymerase II subunit. Mol. Biol. Evolut. 1999, 16, 1799–1808. [Google Scholar] [CrossRef]
O’Donnell, K.; Cigelnik, E. Two divergent intragenomic rDNA ITS2 types within a monophyletic lineage of the fungus Fusarium are nonorthologous. Mol. Phylogenet. Evolut. 1997, 7, 103–116. [Google Scholar] [CrossRef]
Daranagama, D.A.; Camporesi, E.; Tian, Q.; Liu, X.; Chamyuang, S.; Stadler, M.; Hyde, K.D. Anthostomella is polyphyletic comprising several genera in Xylariaceae. Fungal Divers. 2015, 73, 203–238. [Google Scholar] [CrossRef]
Wongkanoun, S.; Becker, K.; Boonmee, K.; Srikitikulchai, P.; Boonyuen, N.; Chainuwong, B.; Luangsa-ard, J.; Stadler, M. Three novel species and a new record of Daldinia (Hypoxylaceae) from Thailand. Mycol. Prog. 2020, 19, 1113–1132. [Google Scholar] [CrossRef]
U’Ren, J.M.; Miadlikowska, J.; Zimmerman, N.B.; Lutzoni, F.; Stajich, J.E.; Arnold, A.E. Contributions of North American endophytes to the phylogeny, ecology, and taxonomy of Xylariaceae (Sordariomycetes, Ascomycota). Mol. Phylogenet. Evolut. 2016, 98, 210–232. [Google Scholar] [CrossRef]
Sir, E.B.; Lambert, C.; Wendt, L.; Hladki, A.I.; Romero, A.I.; Stadler, M. A new species of Daldinia (Xylariaceae) from the Argentine subtropical montane forest. Mycosphere 2016, 7, 596–614. [Google Scholar] [CrossRef]
Miller, M.A.; Pfeiffer, W.; Schwartz, T. Creating the CIPRES science gateway for inference of large phylogenetic trees. In Proceedings of the 2010 Gateway Computing Environments Workshop (GCE), New Orleans, LA, USA, 14 November 2010. [Google Scholar] [CrossRef]
Stamatakis, A. RAxML version 8: A tool for phylogenetic analysis and post–analysis of large phylogenies. Bioinformatics 2014, 30, 1312–1313. [Google Scholar] [CrossRef] [PubMed]
Huelsenbeck, J.P.; Ronquist, F. MrBayes: Bayesian inference of phylogenetic trees. Bioinformatics 2001, 17, 115–755. [Google Scholar] [CrossRef] [PubMed]
Nylander, J.A.A. MrModeltest v. 2.0. Evolutionary Biology Centre; Uppsala University: Uppsala, Sweden, 2004. [Google Scholar]
Swofford, D.L. PAUP*4.0b10: Phylogenetic Analysis Using Parsimony (*and Other Methods); Sinauer: Sunderland, UK, 2002. [Google Scholar] [CrossRef]
Ju, Y.M.; Rogers, J.D. A Revision of the Genus Hypoxylon; Mycologia memoir no.° 20; APS Press: St. Paul, MN, USA, 1996; p. 365. [Google Scholar]
Rogers, J.D.; Ju, Y.M.; Watling, R.; Whalley, A.J.S. A reinterpretation of Daldinia concentrica based upon a recently discovered specimen. Mycotaxon 1999, 72, 507–519. [Google Scholar]
Stadler, M.; Baumgartner, M.; Wollweber, H. Three new Daldinia species with yellowish stromatal pigments. Mycotaxon 2001, 80, 179–196. [Google Scholar]

Figure 1. A maximum likelihood tree of Daldinia based on ITS, LSU, rpb2, and tub2 gene sequences, and CBS 270.87 of Graphostroma platystomum and CBS 12260 of Xylaria hypoxylon as the tree root of Daldinia. The nodes of the branches are labeled MLBS/BPP (MLBS ≥ 70, BPP ≥ 0.9). The known species in this study are shown in blue font, while new species are shown in red font. Ex-type or ex-epitype strains are shown in bold with an “*”. Yellow and green parts are used to distinguish different strains. The black line at the bottom right is the scale bar, indicating 0.1 nucleotide changes at each site.

Figure 2. The distribution and host of Daldinia. (a) the number of Daldinia strains in five provinces of China; (b) the number of families to which the plant hosts of Daldinia.

Table 1. Information on strains used in phylogenetic analysis of the genus Daldinia.

Species	Strains	Country	GenBank Accession Numbers				Reference
Species	Strains	Country	ITS	LSU	rpb2	tub2	Reference
Annulohypoxylon annulatum	CBS 140775	USA	KY610418	KY610418	KY624263	KX376353	[5]
Annulohypoxylon moriforme	CBS 123579	France	KX376321	KY610425	KY624289	KX271261	[5]
Annulohypoxylon nitens	MFLUCC 12.0823	Thailand	KJ934991	KJ934992	KJ934994	KJ934993	[18]
Annulohypoxylon stygium	MUCL 54601	French	KY610409	KY610475	KY624292	KX271263	[5]
Annulohypoxylon truncatum	CBS 140778	USA	KY610419	KY610419	KY624277	KX376352	[5]
Daldinia andina	CBS 114736 *	Ecuador	AM749918	KY610430	KY624239	KC977259	[5]
Daldinia bambusicola	CBS 122872 *	Thailand	KY610385	KY610431	KY624241	AY951688	[5]
Daldinia bambusicola	TBRC 8878	Thailand	MH922869	MH922870	MK165431	MK165422	[7]
Daldinia bambusicola	TBRC 8879	Thailand	MH922872	MH938543	MK165432	MK165423	[7]
Daldinia bambusicola	BCC27937	Thailand	MN153861	MN153876	MN172217	N/A	[19]
Daldinia bambusicola	BCC33678	Thailand	MN153860	MN153877	MN172218	N/A	[19]
Daldinia bambusicola	SAUCC197001	China	PP145311	PP198902	PP263619	PP277065	This study
Daldinia brachysperma	BCC33676	Thailand	MN153854	MN153871	N/A	MN172205	[19]
Daldinia caldariorum	MUCL 49211	France	AM749934	KY610433	KY624242	KC977282	[5]
Daldinia caldariorum	CBS122874	USA	KU683756	KU683756	KU684289	KU684128	[20]
Daldinia chiangdaoensis	BCC88220 *	Thailand	MN153850	MN153867	MN172208	MN172197	[19]
Daldinia chiangdaoensis	BCC88221	Thailand	MN153851	MN153868	MN172209	MN172198	[19]
Daldinia childiae	SAUCC133401	China	PP145313	PP198904	PP263621	PP277067	This study
Daldinia concentrica	CBS 113277	Germany	AY616683	KY610434	KY624243	KC977274	[5]
Daldinia dennisii	CBS 114741 *	Australia	JX658477	KY610435	KY624244	KC977262	[5]
Daldinia eschscholtzii	MUCL 45435	Benin	JX658484	KY610437	KY624246	KC977266	[5]
Daldinia eschscholtzii	TBRC 8876	Thailand	MH938532	MH938541	MK165429	MK165420	[7]
Daldinia eschscholtzii	BCC27887	Thailand	MN153861	MN153878	MN172214	N/A	[19]
Daldinia eschscholtzii	BCC28091	Thailand	MN153862	MN153879	MN172215	N/A	[19]
Daldinia eschscholtzii	BCC62428	Thailand	MN153863	MN153880	MN172216	N/A	[19]
Daldinia eschscholtzii	SAUCC265301	China	PP145315	PP198906	PP263623	PP277069	This study
Daldinia ehretiae	SAUCC228302 *	China	PP145319	PP198888	PP263613	PP277051	This study
Daldinia ehretiae	SAUCC228303	China	PP145320	PP198889	PP263614	PP277052	This study
Daldinia flavogranulata	BCC89363 *	Thailand	MN153856	MN153873	MN172211	MN172200	[19]
Daldinia flavogranulata	BCC89365	Thailand	MN153857	MN153874	MN172212	MN172201	[19]
Daldinia flavogranulata	BCC89376	Thailand	MN153858	MN153875	MN172213	MN172202	[19]
Daldinia jianfengensis	SAUCC373804 *	China	PP145325	PP198890	PP263615	PP277053	This study
Daldinia jianfengensis	SAUCC373805	China	PP145326	PP198891	PP263616	PP277054	This study
Daldinia korfii	EBS 067	Argentina	KY204018	N/A	N/A	KY204014	[21]
Daldinia korfii	EBS 473	Argentina	KY204020	N/A	N/A	KY204016	[21]
Daldinia kretzschmarioides	TBRC 8875	Thailand	MH938531	MH938540	MK165425	MK165416	[7]
Daldinia ledongensis	SAUCC393602 *	China	PP145327	PP198892	N/A	PP277055	This study
Daldinia ledongensis	SAUCC393603	China	PP145328	PP198893	N/A	PP277056	This study
Daldinia loculatoides	CBS 113279	UK	AF176982	KY610438	KY624247	KX271246	[5]
Daldinia macaronesica	CBS 113040	Spain	KY610398	KY610477	KY624294	KX271266	[5]
Daldinia menghaiensis	SAUCC242404 *	China	PP145323	PP198894	PP263617	PP277057	This study
Daldinia menghaiensis	SAUCC242405	China	PP145324	PP198895	PP263618	PP277058	This study
Daldinia phadaengensis	BCC89349 *	Thailand	MN153852	MN153869	MN172206	MN172195	[19]
Daldinia phadaengensis	BCC89350	Thailand	MN153853	MN153870	MN172207	MN172196	[19]
Daldinia petriniae	MUCL 49214	Austria	AM749937	KY610439	KY624248	KC977261	[5]
Daldinia placentiformis	MUCL 47603	Mexico	AM749921	KY610440	KY624249	KC977278	[5]
Daldinia pyrenaica	MUCL 53969	France	KY610413	KY610413	KY624274	KY624312	[5]
Daldinia rhododendri	SAUCC460001 *	China	PP145330	PP198896	N/A	PP277059	This study
Daldinia rhododendri	SAUCC460002	China	PP145329	PP198897	N/A	PP277060	This study
Daldinia spatholobi	SAUCC203501 *	China	PP145318	PP198898	N/A	PP277061	This study
Daldinia spatholobi	SAUCC203502	China	PP145317	PP198899	N/A	PP277062	This study
Daldinia steglichii	MUCL 43512	Papua New Guinea	KY610399	KY610479	KY624250	KX271269	[5]
Daldinia subvernicosa	TBRC 8877 *	Thailand	MH938533	MH938542	MK165430	MK165421	[7]
Daldinia theissenii	CBS 113044	Argentina	KY610388	KY610441	KY624251	KX271247	[5]
Daldinia thunbergiae	SAUCC228601 *	China	PP145322	PP198900	N/A	PP277063	This study
Daldinia thunbergiae	SAUCC228602	China	PP145321	PP198901	N/A	PP277064	This study
Daldinia vernicosa	CBS 119316	Germany	KY610395	KY610442	KY624252	KC977260	[5]
Graphostroma platystomum	CBS 270.87 *	France	JX658535	DQ836906	KY624296	HG934108	[5]
Hypomontagnella monticulosa	MUCL 54604	French	KY610404	KY610487	KY624305	KX271273	[5]
Hypomontagnella monticulosa	BCC58592	Thailand	MN153864	MN153881	MN172219	MN172204	[19]
Hypomontagnella monticulosa	BCC69203	Thailand	MN153865	MN153882	MN172220	MN172203	[19]
Hypomontagnella submonticulosa	CBS 115280	France	KC968923	KY610457	KY624226	KC977267	[5]
Hypoxylon crocopeplum	CBS 119004	France	KC968907	KY610445	KY624255	KC977268	[5]
Hypoxylon fragiforme	MUCL 51264	Germany	KC477229	KM186295	KM186296	KX271282	[5]
Hypoxylon fuscum	CBS 113049	France	KY610401	KY610482	KY624299	KX271271	[5]
Hypoxylon haematostroma	MUCL 53301	France	KC968911	KY610484	KY624301	KC977291	[5]
Hypoxylon haematostroma	BCC50533	Thailand	MN153866	MN153883	MN172221	N/A	[19]
Hypoxylon investiens	CBS 118183	Malaysia	KC968925	KY610450	KY624259	KC977270	[5]
Hypoxylon lateripigmentum	MUCL 53304 *	France	KC968933	KY610486	KY624304	KC977290	[5]
Hypoxylon lenormandii	CBS 119003	Ecuador	KC968943	KY610452	KY624261	KC977273	[5]
Hypoxylon petriniae	CBS 114746 *	France	KY610405	KY610491	KY624279	KX271274	[5]
Hypoxylon rickii	MUCL 53309	France	KC968932	KY610416	KY624281	KC977288	[5]
Hypoxylon rubiginosum	MUCL 52887	Germany	KC477232	KY610469	KY624266	KY624311	[5]
Hypoxylon samuelsii	MUCL 51843	France	KC968916	KY610466	KY624269	KC977286	[5]
Jackrogersella cohaerens	CBS 119126	Germany	KY610396	KY610497	KY624270	KY624314	[5]
Jackrogersella minutella	CBS 119015	Portugal	KY610381	KY610424	KY624235	KX271240	[5]
Jackrogersella multiformis	CBS 119016	Germany	KC477234	KY610473	KY624290	KX271262	[5]
Pyrenopolyporus hunteri	MUCL 52673	Ivory Coast	KY610421	KY610472	KY624309	KU159530	[5]
Pyrenopolyporus laminosus	MUCL 53305 *	France	KC968934	KY610485	KY624303	KC977292	[5]
Pyrenopolyporus laminosus	TBRC 8871	Thailand	MH938527	MH938536	MK165424	MK165415	[7]
Pyrenopolyporus laminosus	BCC89383	Thailand	MN153855	MN153872	MN172210	MN172199	[19]
Pyrenopolyporus nicaraguensis	CBS 117739 *	Burkina Faso	AM749922	KY610489	KY624307	KC977272	[5]
Pyrenopolyporus symphyon	TBRC 8873	Thailand	MH938529	MH938538	MK165428	MK165419	[7]
Xylaria hypoxylon	CBS12260	Sweden	KY610407	KY610495	KY624231	KX271279	[5]

Notes: The new strains for phylogenetic analysis introduced in this experiment are shown in bold. Ex-type or ex-epitype strains are marked with an “*”. N/A: Not available.

Table 2. Collection information of Daldinia specimens.

Species	Collection Location	Collection Time	Strains	Host	Illustration
Daldinia bambusicola	Yunnan Province, Jinghong City, Xishuangbanna primitive forest Park, 22°1′52″ N, 100°52′36″ E	17 March 2023	SAUCC197001	Viburnum rhytidophyllum	Figure 3a
			SAUCC203501	Spatholobus suberectus	Figure 4a
			SAUCC204601	Piper Nigrum	Figure 4b
			SAUCC206901	Cinnamomum verum	Figure 4g
	Yunnan Province, Menghai County, Nanuo Mountain, 21°55′25″ N, 100°35′41″ E	18 March 2023	SAUCC240501	Koelreuteria paniculata	Figure 4c
	Hainan Province, Lingshui County, Diaoluo Mountain, 18°41′45″ N, 109°56′26″ E	8 April 2023	SAUCC283401	Ficus hirta	Figure 4d
	Hainan Province, Ledong County, Jianfengling National Forest Park, 18°41′18″ N, 108°51′31″ E	12 April 2023	SAUCC392801	Schima superba	Figure 4e
	Sichuan Province, Chengdu City, Dujiangyan, 30°59′58″ N, 108°51′31″ E	24 June 2023	SAUCC423801	Citrus maxima	Figure 4f
	Sichuan Province, Leshan City, 29°34′56″ N, 103°17′20″ E	25 June 2023	SAUCC433001	Phyllostachys heteroclada	Figure 4h
	Sichuan Province, Liangshan Prefecture, Xichang City, 27°45′29″ N, 102°18′31″ E	2 July 2023	SAUCC518401	Ageratina adenophora	Figure 4i
	Guizhou Province, Qiandongnan Prefecture, Majiang County, Gudong Town, 26°25′5″ N, 107°21′18″ E	22 August 2023	SAUCC551401	Lophatherum gracile	Figure 4j
	Guizhou Province, Qiannan Prefecture, Sandu County, 25°55′52″ N, 107°57′34″ E	24 August 2023	SAUCC570801	Ulmus pumila	Figure 4k
Daldinia childiae	Fujian Province, Wuyishan City, Xingcun Town, 27°45′5″ N, 117°41′3″ E	15 October 2022	SAUCC133401	Machilus nanmu	Figure 5a
		15 October 2022	SAUCC148701	Eurya japonica	Figure 6a
	Yunnan province, Jinghong City, Xishuangbanna primitive Forest Park, 22°1′52″ N, 100°52′36″ E	17 March 2023	SAUCC209301	Piper nigrum	Figure 6e
	Yunnan Province, Jinghong City, Binjiang Avenue, 22°0′41″ N, 100°48′15″ E	20 March 2023	SAUCC220001	Microstegium vimineum	Figure 6b
	Hainan Province, Lingshui County, Benhao Town, 18°41′54″ N, 109°52′51″ E	9 April 2023	SAUCC314401	Pseudosasa japonica	Figure 6f
	Hainan Province, Ledong County, Jianfengling National Forest Park, 18°42′35″ N, 108°52′35″ E	12 April 2023	SAUCC387901	Litsea cubeba	Figure 6c
			SAUCC388601	Quercus glauca	Figure 6d
			SAUCC392901	Schima superba	Figure 6i
			SAUCC397101	Castanopsis calathiformis	Figure 6g
	Sichuan Province, Leshan City, Erhong Road, 29°35′31″ N, 103°22′39″ E	25 June 2023	SAUCC439301	Symplocos sumuntia	Figure 6h
Daldinia eschscholtzii	Hainan Province, Sanya City, Jiyang District, Dongtian Ridge, 18°23′35″ N, 109°38′14″ E	8 April 2023	SAUCC265301	Lysimachia clethroides	Figure 7a
	Fujian Province, Wuyishan City, Xingcun Town, 27°44′47″ N, 117°40′36″ E	15 October 2022	SAUCC117601	Machilus thunbergii	Figure 8(a1)
			SAUCC123301	Lindera aggregata	Figure 8(a2)
			SAUCC132901	Maesa japonica	Figure 8(a3)
	Yunnan Province, Xishuangbanna Tropical Botanical Garden, 21°55′52″ N, 101°14′50″ E	15 March 2023	SAUCC179601	Gonocaryum lobbianum	Figure 8(b1)
			SAUCC179701	Engelhardia spicata	Figure 8(b2)
			SAUCC182201	Artocarpus heterophyllus	Figure 8(b3)
			SAUCC184501	Tabernaemontana divaricata	Figure 8(b4)
			SAUCC187501	Piper nigrum	Figure 8(b5)
			SAUCC191301	Ficus tinctoria	Figure 8(b6)
			SAUCC191401	Lycianthes biflora	Figure 8(b7)
			SAUCC192301	Bischofia javanica	Figure 8(b8)
	Yunnan province, Jinghong City, Xishuangbanna primitive Forest Park, 22°1′52″ N, 100°52′36″ E	17 March 2023	SAUCC197301	Artocarpus hypargyreus	Figure 8(c1)
			SAUCC201001	Murraya exotica	Figure 8(c2)
			SAUCC205701	Fargesia spathacea	Figure 8(c3)
			SAUCC206001	Zingiber zerumbet	Figure 8(c4)
			SAUCC206301	Microstegium vimineum	Figure 8(c5)
			SAUCC215701	Kadsura longipedunculata	Figure 8(c6)
	Yunnan Province, Xishuangbanna Prefecture, Jinghong City, 22°0′41″ N, 100°48′15″ E	20 March 2023	SAUCC217201	Iris tectorum	Figure 8(d3)
			SAUCC217901	Cordyline fruticosa	Figure 8(d2)
			SAUCC218101	Graptophyllum pictum	Figure 8(d1)
			SAUCC220401	Calliandra haematocephala	Figure 8(d4)
			SAUCC220501	Ficus subulata	Figure 8(d5)
	Yunnan Province, Menghai County, Nanuo Mountain, 21°55′25″ N, 100°35′41″ E	18 March 2023	SAUCC231401	Castanopsis calathiformis	Figure 8(e1)
			SAUCC237801	Betula utilis	Figure 8(e2)
			SAUCC239601	Ageratina adenophora	Figure 8(e3)
			SAUCC243701	Dendrocalamus latiflorus	Figure 8(e4)
			SAUCC244401	Tithonia diversifolia	Figure 8(e5)
	Hainan Province, Lingshui County, Diaoluo Mountain, 18°43′35″ N, 109°52′1″ E	9 April 2023	SAUCC296301	Mangifera indica	Figure 8(f1)
		9 April 2023	SAUCC297601	Lindera nacusua	Figure 8(f2)
	Hainan Province Lingshui County, Benhao town, 18°41′54″ N, 109°52′51″ E	9 April 2023	SUACC300301	Syzygium levinei	Figure 8(g1)
			SAUCC309901	Pseudosasa japonica	Figure 8(g3)
			SAUCC311501	Eurya groffii	Figure 8(g2)
			SAUCC314101	Exbucklandia populnea	Figure 8(g4)
			SAUCC314201	Bridelia balansae	Figure 8(g5)
	Hainan Province, Baoting County, Baocheng town, 109°41′35″ N, 18°42′6″ E	10 April 2023	SAUCC323201	Pronephrium gymnopteridifrons	Figure 8h
	Hainan Province, Changjiang County, Qicha Town, 19°7′2″ N, 109°9′1″ E,	11 April 2023	SAUCC331301	Parashorea chinensis	Figure 8i
	Hainan Province, Changjiang County, Bawangling National Forest Park, 19°7′17″ N, 109°7′6″ E	11 April 2023	SAUCC342001	Clematis uncinata	Figure 8(j1)
			SAUCC343301	Diplospora dubia	Figure 8(j2)
			SAUCC364301	Tilia cordata	Figure 8(j3)
			SAUCC367501	Triadica cochinchinensis	Figure 8(j4)
	Hainan Province, Ledong County, Jianfeng town, 18°42′35″ N, 108°52′35″ E	12 April 2023	SAUCC367701	Rhaphiolepis indica	Figure 8(k1)
			SAUCC369301	Dryopteris podophylla	Figure 8(k2)
			SAUCC371901	Chengiodendron matsumuranum	Figure 8(k3)
			SAUCC373001	Schima superba	Figure 8(k4)
			SAUCC373901	Camellia oleifera	Figure 8(k5)
			SAUCC374001	Polyspora chrysandra	Figure 8(k6)
			SAUCC374501	Lithocarpus henryi	Figure 8(k7)
			SAUCC374601	Eurya nitida	Figure 8(k8)
			SAUCC375901	Elaeocarpus decipiens	Figure 8(k9)
			SAUCC376101	Meliosma rigida	Figure 8(k10)
			SAUCC376901	Rhododendron latoucheae	Figure 8(k11)
			SAUCC377201	Litsea cubeba	Figure 8(k12)
	Hainan Province, Ledong County, Jianfengling National Forest Park, 18°41′18″ N, 108°51′31″ E	12 April 2023	SAUCC394901	Psidium cattleyanum	Figure 8l
	Sichuan Province, Chengdu City, Dujiangyan, 30°59′58″ N, 108°51′31″ E	24 June 2023	SAUCC424501	Phoebe zhennan	Figure 8(m1)
		24 June 2023	SAUCC426101	Rhus chinensis	Figure 8(m2)
	Sichuan Province, Yaan City, Tianquan County, 30°0′21″ N, 102°30′25″ E	26 June 2023	SAUCC440101	Machilus nanmu	Figure 8n
	Guizhou Province, Qiannan Prefecture, Pingtang County, 25°47′42″ N, 107°23′10″ E	23 August 2023	SAUCC561601	Liquidambar formosana	Figure 8o
Daldinia ehretiae	Yunnan Province, Jinghong City, Sancha River, 22°10′10″ N, 100°51′49″ E	19 March 2023	SAUCC228302	Ehretia acuminata	Figure 9a
Daldinia rhododendri	Yunnan Province, Diqing Prefecture, Shangri-la City, 27°58′43″ N, 99°34′24″ E	28 June 2023	SAUCC460001	Rhododendron decorum	Figure 10a
Daldinia spatholobi	Yunnan Province, Jinghong City, Xishuangbanna primitive Forest Park, 22°1′52″ N, 100°52′36″ E	17 March 2023	SAUCC203501	Spatholobus suberectus	Figure 11a
Daldinia thunbergiae	Yunnan Province, Jinghong City, San-cha River, 22°10′10″ N, 100°51′49″ E	19 March 2023	SAUCC228601	Thunbergia grandiflora	Figure 12a
Daldinia jianfengensis	Hainan Province, Ledong County, Jianfeng Town, 18°41′19″ N, 108°51′31″ E	12 April 2023	SAUCC373804	decayed leaves
Daldinia ledongensis	Hainan Province, Ledong County, 18°41′18″ N, 108°51′31″ E	12 April 2023	SAUCC393602	decayed leaves
Daldinia menghaiensis	Yunnan Province, Menghai County, 21°55′25″ N, 100°35′41″ E	18 March 2023	SAUCC242404	decayed leaves

Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

© 2024 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

Share and Cite

MDPI and ACS Style

Yin, C.; Zhang, Z.; Wang, S.; Liu, W.; Zhang, X. A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China. J. Fungi 2024, 10, 700. https://doi.org/10.3390/jof10100700

AMA Style

Yin C, Zhang Z, Wang S, Liu W, Zhang X. A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China. Journal of Fungi. 2024; 10(10):700. https://doi.org/10.3390/jof10100700

Chicago/Turabian Style

Yin, Changzhun, Zhaoxue Zhang, Shi Wang, Wenwen Liu, and Xiuguo Zhang. 2024. "A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China" Journal of Fungi 10, no. 10: 700. https://doi.org/10.3390/jof10100700

APA Style

Yin, C., Zhang, Z., Wang, S., Liu, W., & Zhang, X. (2024). A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China. Journal of Fungi, 10(10), 700. https://doi.org/10.3390/jof10100700

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Menu

A Taxonomic and Phylogenetic Study of Anamorphic Strains of Daldinia (Hypoxylaceae, Xylariales) in Southern China

Abstract

1. Introduction

2. Materials and Methods

2.1. Sample Treatment and Morphological Characterizations

2.2. DNA Extraction and Sequencing

2.3. Phylogenetic Analyses

3. Results

3.1. Phylogenetic Analyses

3.2. Sample Information Statistics

3.3. Taxonomy

4. Discussion

Supplementary Materials

Author Contributions

Funding

Institutional Review Board Statement

Informed Consent Statement

Data Availability Statement

Conflicts of Interest

References

Share and Cite

Article Metrics

Article Access Statistics

Further Information

Guidelines

MDPI Initiatives

Follow MDPI