Taxonomy and Phylogeny of Fungi Associated with Mangifera indica from Yunnan, China

During investigations of saprobic fungi associated with mango (Mangifera indica) in Baoshan and Honghe of Yunnan Province (China), fungal taxa belonging to the orders Botryosphaeriales, Calosphaeriales, Chaetothyriales, Diaporthales, and Xylariales were recorded. Morphological examinations coupled with phylogenetic analyses of multigene sequences (ITS, LSU, SSU, tef1-α, rpb1, rpb2, β-tubulin and CAL) were used to identify the fungal taxa. A new genus viz. Mangifericola, four new species viz. Cyphellophora hongheensis, Diaporthe hongheensis, Hypoxylon hongheensis, and Mangifericola hongheensis, four new host and geographical records viz. Aplosporella artocarpi, Hypomontagnella monticulosa, Paraeutypella citricola and Pleurostoma ootheca, and two new collections of Lasiodiplodia are reported.


Introduction
Mango (Mangifera indica L.) is a dicotyledonous fruit plant in the family Anacardiaceae, and the genus Mangifera contains approximately 69 species with more than 1000 varieties [1]. Mango cultivation history can be traced back 4000 years in India and Southeast Asia. As one of the five most economically significant fruit crops worldwide, Mango is cultivated in more than 100 countries, of them, Asian countries account for approximately 77% of the world's mango production [2][3][4]. India is the largest mango producer, accounting for about 54.2% of the total mangoes produced worldwide, while China, Thailand, Indonesia, Mexico, and Pakistan are other major mango producers [2][3][4]. Mango planting is an important part of agricultural exports in sub-tropical to tropical countries [3][4][5]. Naturally, mango trees grow best in lowland subtropical to tropical regions, best in dry, sandy soil with a pH of 5.5-7.5, and direct sun is preferred for tree growth and fruit production [6].
The investigation of plant-associated microfungi is important, as is relevant to the trend towards globalization of agricultural markets, including forest and horticultural Box (8.2.4) [36,37] in the CIPRES Science Gateway v.3.3 (http://www.phylo.org/portal2, accessed on 2 May 2022 [38]) with GTRGAMMA substitution model with 1000 bootstrap iterations. The Bayesian analysis was performed by MrBayes on XSEDE (3.2.7a) via the CIPRES Science Gateway V.3.3 web server [38]. Bayesian posterior probabilities (BYPP) [39] were evaluated by Markov Chain Monte Carlo sampling (MCMC). The best models of evolution were estimated by using MrModeltest v. 2.3 [40] and PAUP v. 4.0b10 [41]. Six simultaneous Markov chains were run for 1,000,000 to 10,000,000 generations, depending on individual settings for the fungal group and the resulting trees were sampled at one tree every 1000th generation. Phylogenetic trees were visualized using FigTree v1.4.0 [42] and the trees were edited by Microsoft PowerPoint and inserted with reliable statistical supports from ML and BI. Type species: Mangifericola hongheensis E.F. Yang & Tibpromma Saprobic on dead terricolous wood. Sexual morph: Pseudostroma poorly developed, delimited with a black surface, irregular, and raised. Ascomata immersed, irregular to subglobose, clustered, visible emerging apical parts of extended neck on distinct black region. Ostioles cylindrical, sulcate, ostiolar canal periphysate. Peridium thick near ostiole, multilayers, dark brown-walled outwardly, comprised by hyaline, compressed textura angularis cells to inner layers. Paraphyses not seen. Asci 8-spored, cylindrical to oblong, unitunicate, pedicellate, straight to fairly curved, hyaline, apically flat. Ascospores oblong to allantoid, hyaline, or yellowish, with oil droplets at both ends when mature. Asexual morph: Undetermined.

Taxonomy and Phylogenetic Analyses Results
Notes: Mangifericola (M.) generally fits with the common concept of Diatrypaceae by having ascomata immersed, subglobose or irregular, with elongated neck, hamathecium absent, asci eight-spored, straight to fairly curved, allantoid ascospores, hyaline or yellowish, with oil droplets [43,44]. However, Mangifericola is mainly distinguished from other genera in this family by having a single prolonged neck erect from black pigmented lesions on the wood surface. Additionally, this new genus is distinguished from other closely related genera based on phylogenetic analyses ( Figure 1). The results from BLASTn searches of ITS, LSU, and β-tubulin are shown in Table 1. Despite the BLASTn results of ribosomal DNA regions of Mangifericola indicating it is closely related to Diatrype and Diatrypella, the β-tubulin gene indicated it is related to Melanostictus with a low similarity. Diatrypella is characterized by cushion-like or discoid stromata, umbilicate or sulcate ostiolar necks, cylindrical, polysporous, long-stalked asci, and allantoid, hyaline or yellowish ascospores [45]. Our new genus is similar to Diatrype by having allantoid, hyaline or yellowish ascospores but differs in having poorly developed pseudostroma, irregular to subglobose ascomata with a single prolonged neck erect from black pigmented and cylindrical to oblong asci. Therefore, we establish Mangifericola (Diatrypaceae) as a distinct new genus.
subglobose ascomata with a single prolonged neck erect from black pigmented and cylindrical to oblong asci. Therefore, we establish Mangifericola (Diatrypaceae) as a distinct new genus.  Phylogram of a novel genus Mangifericola, a new host record of Paraeutypella citricola, and other genera within the family Diatrypaceae generated from maximum likelihood analysis based on a combined ITS, β-tubulin sequence datasets, with Xylaria berteroi (YMJ 95101511) and Kretzschmaria deusta (CBS 826.72) as the outgroups. Related sequences used in the phylogeny were taken from Dissanayake et al. [26]. The species introduced in this study are indicated in red, and the type strains are indicated in bold with "T". Bootstrap values equal to or greater than 70% (ML, Left) and Bayesian posterior probabilities (BI, right) equal to or greater than 0.95 are given at the nodes. Hyphens (-) represent values less than 70% in ML/0.95 in BI. For more information, please see supplementary materials (Table S1, Supplementary Information S1). Phylogram of a novel genus Mangifericola, a new host record of Paraeutypella citricola, and other genera within the family Diatrypaceae generated from maximum likelihood analysis based on a combined ITS, β-tubulin sequence datasets, with Xylaria berteroi (YMJ 95101511) and Kretzschmaria deusta (CBS 826.72) as the outgroups. Related sequences used in the phylogeny were taken from Dissanayake et al. [26]. The species introduced in this study are indicated in red, and the type strains are indicated in bold with "T". Bootstrap values equal to or greater than 70% (ML, Left) and Bayesian posterior probabilities (BI, right) equal to or greater than 0.95 are given at the nodes. Hyphens (-) represent values less than 70% in ML/0.95 in BI. For more information, please see Supplementary Materials (Table S1, Supplementary Information S1).  Fungal Name number: FN 571237 Etymology: The name reflects the location "Honghe" where the holotype was collected.
Holotype: HKAS 122665 Saprobic on dead branch of Mangifera indica. Sexual morph: Pseudostroma poorly developed, delimited with a black surface, irregular, and raised. Ascomata (exclude neck)  Notes: Mangifericola hongheensis was collected from a dead wood piece of Mangifera indica, and it clearly differs from other taxa in the family Diatrypaceae based on multigene phylogenetic analyses and morphological comparisons. The new species formed a well-separated clade distant from other genera within Diatrypaceae. Morphologically, our species is relatively similar to Diatrype palmicola (MFLU 15-0040, holotype), and they all cause black pigmented lesions on the wood epidermis, ascomata clustered as small groups, fully immersed, visible extended neck raised above, and the absence of hamathecium ( Figure 2). However, the ascomata are different by coloration (dark black vs. brown), and the asci of our species have a shorter pedicel [46]. The comparison of the ITS, and LSU bp regions also showed that the two species have big differences in base pairs (ITS: 8.1%; LSU: 3.1% bp differences) [46], but we were not able to compare the β-tubulin gene of D. palmicola as they lack of β-tubulin gene. In addition, we compared the M. hongheensis with the closely related species in phylogeny and from BLASTn results (Melanostictus sp., Pedumispora sp., Halodiatrype sp., Diatrypella sp. and Diatrype sp.) but they differ. Therefore, based on both unique morphological characteristics and molecular data we establish  [47] with the type species Hypoxylon fragiforme (Pers.). The generic concept of Hypoxylon traditionally differs from other genera in the family Xylariaceae by four main characteristics viz. Nodulisporium-like asexual morph; unipartite stromata; solid and homogeneous stromatal tissue below the perithecial layer; and stromata not upright [48][49][50]. Later, the molecular studies led to the segregation of further genera such as; Annulohypoxylon [51], Hypomontagnella [52], Jackrogersella, and Pyrenopolyporus [53] which were previously considered Hypoxylon taxa. Hypoxylon as the type genus accommodates 829 records in Index Fungorum [16]. The members of Hypoxylon frequently grow on dead wood as wood degrading fungi, and they are also often isolated as endophytes of seed plants [52,53]. Hypoxylon species are an excellent source of bioactive secondary metabolites, e.g., H. fuscum [54]. The phylogenetic relationships of this generic species are shown in Figure 3.   Likelihood analysis based on a combined LSU, β-tubulin, ITS and rpb2 sequence datasets, with Biscogniauxia nummularia (MUCL 51395) and Xylaria hypoxylon (CBS 122620) as the outgroups. Related sequences used in the phylogeny were taken from Song et al. [29]. The species introduced in this study are indicated in red, and the type strains are indicated in bold with "T". Bootstrap values equal to or greater than 70% (ML, Left) and Bayesian posterior probabilities (BI, right) equal to or greater than 0.95 are given at the nodes. Hyphens (-) represent values less than 70% in ML/0.95 in BI. For more information, please see supplementary materials (Table S2, Supplementary  Information S2).
Holotype: HKAS 122663 . Phylogram of a new species Hypoxylon hongheensis, a new host record of Hypomontagnella monticulosa, and related genera within the family Hypoxylaceae generated from Maximum Likelihood analysis based on a combined LSU, β-tubulin, ITS and rpb2 sequence datasets, with Biscogniauxia nummularia (MUCL 51395) and Xylaria hypoxylon (CBS 122620) as the outgroups. Related sequences used in the phylogeny were taken from Song et al. [29]. The species introduced in this study are indicated in red, and the type strains are indicated in bold with "T". Bootstrap values equal to or greater than 70% (ML, Left) and Bayesian posterior probabilities (BI, right) equal to or greater than 0.95 are given at the nodes. Hyphens (-) represent values less than 70% in ML/0.95 in BI. For more information, please see Supplementary Materials (Table S2, Supplementary Information S2).
Culture characteristics: Colonies on PDA 50 mm in diameter after two weeks at 27 • C, gray, after around two months, colonies on PDA becoming reddish-brown, circular, regular margin, flat; dark brown at the reverse, with reddish pigments produced in PDA. No sporulation on PDA and oatmeal agar (OMA) media within three months, Vegetative hyphae 2-4 µm wide, hyaline, smooth-walled. Notes: Based on morphology, our isolates fit with the concept of Hypoxylon by having effused-pulvinate, unipartite ascomata, with solid and homogeneous, stromatal tissue. The BLASTn results for ITS, rpb2, β-tubulin, and LSU region are shown in Table 1. In addition, the phylogenetic analysis of combined LSU, ITS, rpb2 and β-tubulin sequence showed our strains (HKAS 122663, KUMCC 21-0452) separate well from H. perforatum (CBS 115281) ( Figure 3). Following the description of H. perforatum by Khodaparast [55] and Kout [57] with the type species D. eres, and it was placed in the family Diaporthaceae [58]. Previously, taxa in this genus were known as the asexual morph and named as Phomopsis, but it was replaced by the sexual morph typified name Diaporthe [59]. The species identification of Diaporthe spp. is traditionally based on host association and phenotypic features, and the sexual morph is characterized by having immersed ascomata, with erumpent pseudostroma, fusoid, ellipsoid to cylindrical, hyaline ascospores, with or without septate, and sometimes having appendages [57,60], while the asexual morph produces three kinds of conidia viz. α-conidia (straight, guttulate or eguttulate, smooth-walled), β-conidia (straight or hamate, smooth-walled, guttulate), and γ-conidia (seldom produced, multiguttulate, fusiform to subcylindrical, apically acute or rounded while the base is sometimes truncate) [61]. Diaporthe species have been reported occurring on various plants as pathogens, saprobes, or endophytes worldwide, and in addition, Diaporthe is responsible for many diseases such as root and fruit rot, dieback, cankers, leaf spot, leaf and pod blights, wilt, and seed decay of economically important agricultural crops or woody hosts [62]. Additionally, Diaporthe species have been reported as pathogens associated with humans and other mammals [63]. Diaporthe also has the potential to stop herbivory by lignocellulolytic activities [64], and Ash et al. [65] reported its use as a bioherbicide. The phylogeny of this genus is shown in Figure 5. α-conidia (straight, guttulate or eguttulate, smooth-walled), β-conidia (straight or hamate, smooth-walled, guttulate), and γ-conidia (seldom produced, multiguttulate, fusiform to subcylindrical, apically acute or rounded while the base is sometimes truncate) [61]. Diaporthe species have been reported occurring on various plants as pathogens, saprobes, or endophytes worldwide, and in addition, Diaporthe is responsible for many diseases such as root and fruit rot, dieback, cankers, leaf spot, leaf and pod blights, wilt, and seed decay of economically important agricultural crops or woody hosts [62]. Additionally, Diaporthe species have been reported as pathogens associated with humans and other mammals [63]. Diaporthe also has the potential to stop herbivory by lignocellulolytic activities [64], and Ash et al. [65] reported its use as a bioherbicide. The phylogeny of this genus is shown in Figure 5.    (Table S3, Supplementary Information S3).
Culture characteristics: Colonies on PDA 50-70 mm in diameter after two weeks at 27 °C, white to gray, circular, flat to effuse, medium dense, fimbriate margin; white at the reverse, without pigments produced in PDA, but produced black dots and released fluid secretions after one month, but without any spores were observed. Vegetative hyphae 1-3 µm wide, hyaline, smooth-walled. Figure 5. Phylogram of a new species Diaporthe hongheensis and closed species within genus Diaporthe generated from maximum likelihood analysis based on a combined ITS, tef1-α, β-tubulin and CAL sequence datasets, with Diaporthella corylina (CBS 121124) as the outgroup. Related sequences used in the phylogeny were taken from Ariyawansa et al. [66] and Dong et al. [28]. The species introduced in this study are indicated in red, and the type strains are indicated in bold with "T". Bootstrap values equal to or greater than 70% (ML, Left) and Bayesian posterior probabilities (BI, right) equal to or greater than 0.95 are given at the nodes. Hyphens (-) represent values less than 70% in ML/0.95 in BI. For more information, please see the Supplementary Materials (Table S3, Supplementary Information S3).
Culture characteristics: Colonies on PDA 50-70 mm in diameter after two weeks at 27 • C, white to gray, circular, flat to effuse, medium dense, fimbriate margin; white at the reverse, without pigments produced in PDA, but produced black dots and released fluid secretions after one month, but without any spores were observed. Vegetative hyphae 1-3 µm wide, hyaline, smooth-walled.  Notes: Diaporthe pseudomangiferae and D. pseudophoenicicola were isolated from Mangifera indica with or without pathogenetic symptoms [63]. Our new isolate D. hongheensis fits well with the concept of Diaporthe by having fully immersed, mostly subglobose, ellipsoid to cylindrical ascomata, hyaline, septate ascospores with appendages [58]. Diaporthe hongheensis is different by having 6(-8)-spores asci, while most Diaporthe species which available sexual morph have 8-spored asci, like D. eucalyptorum, D. alnea, and D. neilliae [67]. The BLASTn results of ITS, β-tubulin, CAL, and tef1-α gene region were shown in Table 1. In addition, the multi-locus phylogenetic analysis separated D. hongheensis (KUMCC 21-0457) well although closely related to D. viniferae, D. pandanicola and D. fraxini-angustifoliae clades. In addition, for those phylogenetically closely related species, only asexual morphs are available. Therefore, we introduce D. hongheensis as novel species based on morphomolecular analyses.
Eurotiomycetes  16: 47 (1962) Notes: The genus Cyphellophora (C.) with C. laciniata as the type species was established in 1962 [68]. To date, Cyphellophora contains a total of 31 species excluding three species that have been transferred to Aphanophora, Camptophora, and Pseudomicrodochium [46]. The members of Cyphellophora species are usually reported as saprobic on excretions of insects or foliar epiphytes on living leaves worldwide, and also as endophytic fungi on fresh leaves, while Yang et al. [69] introduced the first sexual morph record with the new species C. jingdongensi (IFRD 9049) from living leaves of Alnus nepalensis in China. In addition, C. laciniata, C. europaea and C. pluriseptata were found associated with human and animal skin and nails [70]. Cyphellophora sessilis was reported as a pathogenic fungus that causes sooty blotch, flyspeck, and diseases of certain fruit corps [71]. The sexual morph of Cyphellophora is characterized by ascomata fusing with host tissue at the base, scattered, subglobose to globose, dark brown, and ostiole inconspicuous; asci each ellipsoidal to cylindrical, short pedicel, bitunicate, hyaline, and septate ascospores. Based on a previous study, the asexual morph of this genus was described by producing branched hyphae, intercalary, terminal, or lateral, sparse, or integrated; conidiogenous cells phialidic, hyaline or pale brown, conidia ranging in shape from oblong to fusiform or vermiform [70]. The phylogeny of this genus is shown in Figure 7.
Culture characteristics: Colonies on PDA 15-20 mm in diameter after two weeks at 27 • C, brown to black, effuse, circular, dense and rough at the surface, well-defined, undulate edge with sinking, slightly striated; dark brown at the reverse, without pigments produced in PDA. Vegetative hyphae 2-4 µm wide, hyaline, branched, septate.
Material Notes: Our isolates clustered within Cyphellophora, while they share similar characteristics with C. jingdongensi (IFRD 9049) which was reported by Yang et al. [69] from China, but our strains differ by having distinct setae at the outermost of peridium, and ellipsoid to irregular, 1-septate, thick-walled, rough, and smaller (12-16 × 3-5 µm vs. 16-24 × 5-7 µm) ascospores. The BLASTn values and percent-sequence of ITS, SSU, LSU and rpb1 showed that our strain is closely related to the taxa in Table 1, and phylogenetic results indicated that they are separated. Phylogenetic results indicated that our isolates are well separated from C. attinorum (CBS 131958), C. sessilis (CBS 238.93, CBS 243.85) and C. jingdongensi (IFRD 9049) clades with high statistical supports (92% in ML, 0.99 in BI) ( Figure 7). Therefore, based on the evidence of morphology and phylogeny, we introduce our strains as a new species, C. hongheensis on Mangifera indica from China.
relatively wider at upper than lower, guttulate, without a mucilaginous sheath. Asexual morph: Undetermined.
Culture characteristics: Colonies on PDA 15-20 mm in diameter after two weeks at 27 °C, brown to black, effuse, circular, dense and rough at the surface, well-defined, undulate edge with sinking, slightly striated; dark brown at the reverse, without pigments produced in PDA. Vegetative hyphae 2-4 µm wide, hyaline, branched, septate.
Material Notes: Our isolates clustered within Cyphellophora, while they share similar characteristics with C. jingdongensi (IFRD 9049) which was reported by Yang et al. [69] from China, but our strains differ by having distinct setae at the outermost of peridium, and ellipsoid to irregular, 1-septate, thick-walled, rough, and smaller (12-16 × 3-5 µm vs. 16-24 × 5-7 µm) ascospores. The BLASTn values and percent-sequence of ITS, SSU, LSU and rpb1 showed that our strain is closely related to the taxa in Table 1, and phylogenetic results indicated that they are separated. Phylogenetic results indicated that our isolates are well separated from C. attinorum (CBS 131958), C. sessilis (CBS 238.93, CBS 243.85) and C. jingdongensi (IFRD 9049) clades with high statistical supports (92% in ML, 0.99 in BI) ( Figure 7). Therefore, based on the evidence of morphology and phylogeny, we introduce our strains as a new species, C. hongheensis on Mangifera indica from China.     [72] with L. tubericola as the type species, and to date, this genus contains 69 records in Index Fungorum [16]. Sexual morph of Lasiodiplodia species have been poorly documented, but the known sexual morph of L. gonubiensis, L. lignicola, L. theobromae, and L. pseudotheobromae are characterized by producing ellipsoidal to fusiform, aseptate, straight to curved ascospores, hyaline to brown when mature [73]. The asexual morph of Lasiodiplodia was characterized by having pycnidial paraphyses and longitudinal striations on the mature conidia, but identification of Lasiodiplodia spp. only relying on main morphological features (conidia and paraphyses) is impossible. Thus, ITS and tef1-α regions have been widely used to distinguish different species in this genus [74]. This genus is commonly distributed in subtropical to tropical regions where the temperature is high and is associated with various diseases of woody hosts such as stem blight and/or canker, and dieback [74]. Especially, some Lasiodiplodia spp. (L. iraniensis, L. theobromae, and L. laeliocattleyae) and other species in Botryosphaeriaceae are well-known pathogens associated with mango [75,76]. The phylogeny of Lasiodiplodia and closely related genera are shown in Figure 9. Notes: Species of Lasiodiplodia are mostly distinguished by the morphology of the conidia and paraphyses [74], and our isolate L. theobromae fits with the genetic concept of Lasiodiplodia by having hyaline to brown, thick-walled conidia, and pycnidial paraphyses. The BLASTn results of tef1-α, rpb2, ITS, β-tubulin and LSU showed that our isolate is closely related to L. theobromae strains with relatively high similarity (100%) ( Table 1). In addition, the multi-gene phylogenetic analysis also showed our strains (HKAS 122660 and HKAS 122659) were relatively closely related to L. theobromae (CBS 111530, CBS 339.90, and CBS 146.96) (Figure 9). Therefore, L. theobromae is reported as an extra collection from China and associated with mango. addition, the multi-gene phylogenetic analysis also showed our strains (HKAS 122660 and HKAS 122659) were relatively closely related to L. theobromae (CBS 111530, CBS 339.90, and CBS 146.96) (Figure 9). Therefore, L. theobromae is reported as an extra collection from China and associated with mango.     immersed to totally immersed, ampulliform, solitary to gregarious, dark brown, short ostiole, visible apical black region raised on the top surface. Ostioles 43-54 × 57-85 µm (x = 48 × 71 µm, n = 10), single, cylindrical with conical at the apical, brown. Pycnidial walls 23-55 µm (x = 39 µm, n = 20) wide, thick-walled, unequal thickness, the outer layers comprised by brown to dark brown cells of textura globulosa, the inner layers comprised of hyaline to brownish cells of textura angularis. Hamathecium 3-4 µm (x = 3.5 µm, n = 20) wide, cylindrical, hyaline, aseptate, wide at the base, unbranched, round at tip, raised among conidia, paraphyses. Conidiophores reduced to conidiogenous cells. Conidiogenous cells 14-21 × 3-5 µm (x = 17 × 4 µm, n = 20), holoblastic, cylindrical to subcylindrical, hyaline, granules, thick-walled, some slightly bent, producing a single conidium at the top. Conidia 22-28 × 9-14 µm (x = 25 × 11 µm n = 20), obovoid to ellipsoid, hyaline, rounded at the apex, some constricted in the middle, verruculose, granules, thick-walled, wall < 2 µm, hyaline, aseptate, without longitudinal striations or mucilaginous sheath.
Culture characteristics: Colonies on PDA 20 mm in diameter after one week at 25 • C in natural light, circular, flat to effuse, superficial, entire edge, white to gray when its young, hyphae green to brown at the centre, raised hyphal mass after one month, conidia formation in cultures after half year; dark brown to black at the reverse, without pigments produced in PDA. Vegetative hyphae 1-3 µm wide, hyaline, septate, and conidia are same to above descriptions.
Culture characteristics: Colonies on PDA 40 mm in diameter after two weeks at 27 • C. Colonies white, flat, cottony, with a fimbriate edge, with a medium density; grayish at the reverse, without pigments produced in PDA. Vegetative hyphae 1-3 µm wide, hyaline, septated, branched.
Calosphaeriales M.E. Barr [108] with Pl. candollei as the type species. It currently accommodates a total of five species in Index Fungorum [16], and most species have molecular data in GenBank (2022) (https://www.ncbi. nlm.nih.gov/nuccore/, accessed on 4 May 2022). Two Pleurostoma (Pl. vibratile and Pl. minimum) have been reidentified based on phylogenetic analyses and transferred to Phaeoacremonium (Togniniaceae) [109]. Sexual morph of Pleurostoma is characterized by having semi-immersed to erumpent, superficial ascomata, hamathecium absent, obovoid, club-like, pedicellate asci, ascospore numerous, allantoid, aseptate, hyaline, extremely flexuous [110,111]. In addition, the asexual morphologies were mentioned by Tsang et al. and Huang et al. [32,112] as hyphomycetous, with hyaline to brown, branched, septate mycelia, projection-like conidiophores, phialides monophialidic or polyphialidic, cylindrical, erect, straight to fexuouse, hyaline to brown usually laterally located, clustered, having produced hyaline, ovoid to suballantoid, aseptate, smooth-walled conidia on the apex of the phialides, present as a slimy mass. In nature, Pleurostoma is widely distributed in woods, soil, and sewage worldwide, and was mostly reported in Iran, Spain, and Sri Lanka [8]. In addition, Tsang et al. [32] first reported one human-infected case associated with a dematiaceous fungus Pl. hongkongense, which was isolated from the subhepatic abscess pus and drain fluids of a patient. The phylogenetic relationships among the taxa in this genus were well-studied by Tsang et al. [32], and the updated phylogenetic tree of this study is shown in Figure 16. is widely distributed in woods, soil, and sewage worldwide, and was mostly reported in Iran, Spain, and Sri Lanka [8]. In addition, Tsang et al. [32] first reported one humaninfected case associated with a dematiaceous fungus Pl. hongkongense, which was isolated from the subhepatic abscess pus and drain fluids of a patient. The phylogenetic relationships among the taxa in this genus were well-studied by Tsang et al. [32], and the updated phylogenetic tree of this study is shown in Figure 16.     , subglobose to ampulliform, solitary to gregarious, semi-immersed to erumpent, black, conspicuous at the surface, coriaceous, shiny, carbonaceous, ostiolar, without papillate. Ostioles canal 40-55 × 50-65 µm (x = 47 × 57 µm, n = 20), narrowly, mostly in central, brown to black. Peridium 12-16 µm wide, thin-walled, unequal in thickness, thicken near the neck, muti-layers, comprised of compressed textura prismatica, hyaline cells at the inner layers 8-13 µm, membranaceous, and composed of pale brown to dark brown cells of textura intricata to textura epidermoidea

Discussion
To date, around 160 records of mango-associated xylarialean fungi have been documented in the U.S. National Fungus Collections Fungal Database [16]. This study, a novel genus Mangifericola (Diatrypaceae) is established with M. hongheensis as the type species. Based on the BLASTn results of ITS, and LSU, our new genus Mangifericola is closely related to Diatrype and Diatrypella, but the BLASTn result of tub2 indicated that it is a distinct genus (Table 1). In addition, Mangifericola has the characteristics of having ascomata groups fully immersed with a long and erect neck, and the colony characteristics are similar to the Diatrype palmicola (MFLU 15-0040) [46], but they are distinguished by ITS, tub2 gene regions and phylogenetic analyses (Figure 1). The new host record of Paraeutypella citricola (Diatrypaceae) is also introduced in this study based on morphology and multigene phylogeny. The sizes, asci, and ascospore morphology of our strain well matched with previous studies of P. citricola [46,107], and the BLASTn results of ITS, LSU, and tub2 also showed a high similarity (>99%) with P. citricola ( Table 1). The BLASTn results of Hypoxylon hongheensis (Hypoxylaceae) indicated that it is closely related to sister species H. perforatum, however, they differ due to the base pair differences of ITS, rpb2, tub2 (Table 1), asci size and ascomatal pigments in 10% KOH solution [55,56]. Another new host record of a xylarialean fungus, Hypomontagnella monticulosa (Hypoxylaceae) is also described here based on morphology and multigene phylogeny ( Figure 3).
Many species in the Botryosphaeriales have been reported on mango, of which two species belong to the Aplosporellaceae, while 60 species belong to the Botryosphaeriaceae [16]. Aplosporella artocarpi (Aplosporellaceae) is introduced as a new host and country record in this study. As the LSU of A. artocarpi (CPC 22791) and tef1-α of A. chromolaenae (MFLUCC 17-1517) were not available, thus morphological comparisons (Table 2) are provided to support our isolate is Aplosporella artocarpi. We also report two other new collections of Lasiodiplodia theobromae and L. pseudotheobromae (Botryosphaeriaceae), and they all have previously been isolated from mango in China [78,79,85]. The mature conidia of Lasiodiplodia pseudotheobromae (HKAS 122658) and L. theobromae (HKAS 122660) were not found, but the BLASTn results, conidia morphology, and phylogenetic analyses result fully supported the identification.
From the order Calosphaeriales, only Calosphaeria mangiferae (Calosphaeriaceae) has earlier been reported on Mangifera indica [16]. The Pleurostoma (Pleurostomataceae) are widely distributed in woods, soil, and sewage worldwide, and in addition, a dematiaceous fungus Pl. hongkongense was isolated from a patient [32]. In total, 73 records of mango-associated diaporthalesan fungi have been documented in the U.S. National Fungus Collections Fungal Database, of which 78% of the records (57 records) belong to Diaporthe and Phomopsis (Diaporthaceae) [16]. Diaporthe species reported were plant pathogens, saprobes, endophytes, or associated with humans and other mammals [78,87,88]. Our isolate Diaporthe hongheensis (HKAS 122657) is highly matched to the concept of sexual Diaporthe [29,78,84], and based on the BLASTn results and phylogenetic analyses support, we identified it as a new species (Table 1, Figure 5). A chaetothyrialean fungus viz. Cyphellophora hongheensis is reported associated with mango for the first time. Species of Cyphellophora (Cyphellophoraceae, Chaetothyriale) are epiphytic on excretions of insects or foliar epiphytes on living leaves, and some species are associated with human and animal skin and nails, including Cyphellophora laciniata, C. europaea, and C. pluriseptata [71,100]. In this study, our saprobic fungal isolate C. hongheensis was isolated from a living mango branch hanging on a mango tree and is characterized by ascomata that are superficial, and the absence of distinct pathogenetic symptoms.
In this study, we isolated four species of xylarialean fungi, three species of botryosphaerialen fungi, and three species from each order of Calosphaeriales, Chaetothyriales and Diaporthales. Based on the reports of the previous studies and this study, the species of Xylariales and Botryosphaeriales indeed have a high association with mango. Moreover, Cyphellophora, Diaporthe and Pleurostoma seem to have a wide range of adaptions for different hosts viz. plants, human and animals.
Supplementary Materials: The following supporting information can be downloaded at: https: //www.mdpi.com/article/10.3390/jof8121249/s1, Tables S1-S7: The names, isolate numbers, and corresponding GenBank accession numbers of the taxa used in phylogenetic trees; Supplementary Information (S1-S7): the detail information of phylogenetic trees.